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Optimization of a Manufacturing Process of Parenteral
Lyophilised Drugs: Process Validation
Cláudia Rego Rodrigues
Thesis to obtain the Master of Science Degree in
Pharmaceutical Engineering
Supervisors: Eng. Samuel Mendes Geraldes Camocho
Prof. Carlos Manuel Faria de Barros Henriques
Examination Committee
Chairperson: Prof. José Monteiro Cardoso de Menezes
Supervisor: Eng. Samuel Mendes Geraldes Camocho
Member of the Committee: Prof. João Fernandes de Abreu Pinto
November 2019
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I declare that this document is an original work of my own authorship and that it fulfils all the
requirements of the Code of Conduct and Good Practices of the Universidade de Lisboa
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Preface
The work presented in this thesis was performed at department of Liquid an Injectables Division of
Hikma Farmacêutica S.A (Terrugem, Portugal), during the period February – July 2019, under the
supervision of Eng. Samuel Mendes Geraldes Camocho. The thesis was co-supervised at Instituto
Superior Técnico by Prof. Prof. Carlos Manuel Faria de Barros Henriques.
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Abstract
Exhibit batches were manufactured in 2011 and an ANDA with two presentations Product Y for
Injection 5 g/vial Product Y for Injection 10 g/vial were submitted to authorities. The approval was
guaranteed to Hikma by FDA on October 2018.
During launching batches of Product Y 5 g/vial it was noticed a high breakage rate, around 13,6%,
representing a non-robust and not validated process. The improvement of Product Y 5 g/vial was
essential and for that it was necessary to investigate the life cycle of Product Y. During the
investigation it was concluded that the root cause of vial breakage was attributed to the fast-freezing
phase. The fast freezing associated with the high filling volume caused a rapid and large expansion of
the cake resulting in breakage problems.
In order to correct this problem, a Risk Assessment for the lyophilisation cycle between Product Y 5
g/vial and 10 g/vial was made. The lyophilisation cycle and the recipe of Product Y 10 g/vial was then
adapted to the 5 g/vial resulting in a decrease to 0,7% of rejected vials resulting in the product’s
validation. After correction the product was successfully launched.
Additionally, process improvement for efficiency purpose proposed a Line Transfer and a scale-up. For
that, a Process Validation Protocol was designed and during these activities a decrease to 0,4% of
broken vials was observed. With these results it was possible to confirm that our modifications were
made successfully resulting in a robust process and independent from the line of the production.
Keywords: Lyophilisation; Process Validation; Life Cycle; Optimization; Technology Transfer; Scale-
up.
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Resumo
Em 2011, após os lotes de exibição, um ANDA com duas apresentações Produto Y para Injeção 5 g/
frasco Produto Y para Injeção10 g / frasco foram submetidos às autoridades. A aprovação foi
acreditada à Hikma pelo FDA em Outubro de 2018.
Durante o lançamento de lotes do Produto Y 5 g / frasco, verificou-se uma percentagem elevada de
frascos rejeitados, cerca de 13,6%, representando um processo não robusto e, portanto, não
validado. A otimização do Produto Y 5 g / frasco foi essencial e, para isso, investigou-se o ciclo de
vida do produto. Durante a investigação, concluiu-se que a principal causa de frascos partidos deve-
se ao rápido congelamento. O congelamento rápido associado a um volume de enchimento alto
provoca uma expansão rápida do produto, resultando em problemas de quebra.
Para corrigir este problema, foi feita uma Avaliação de Risco para o ciclo de liofilização entre o
Produto Y 5 g / frasco e 10 g / frasco. O ciclo de liofilização do Produto Y 10 g / frasco foi adaptado
para a receita de 5 g / frasco, resultando numa diminuição de frascos rejeitados para 0,7% tornando o
processo validado.
Adicionalmente, uma Transferência de Linha e um “scale-up” foram propostos. Para tal, foi elaborado
um Protocolo de Validação de Processo e, durante as atividades, foi observada uma redução para
0,4% de frascos partidos. Com os resultados obtidos, foi possível confirmar que as modificações
foram feitas com sucesso, resultando num processo robusto e independente da linha de produção.
Palavras-chave: Liofilização; Validação de Processo; Ciclo de Vida; Otimização; Transferência de
Tecnologia; “Scale-up”.
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Acknowledgments
A tese não é apenas resultado de um empenho individual, mas sim de um conjunto de esforços que o
tornaram possível e sem os quais teria sido muito mais difícil chegar ao fim desta etapa, que
representa um importante marco na minha vida pessoal e profissional. Desta forma, manifesto a
minha gratidão a todos os que estiveram presentes no decorrer destes últimos anos.
Em primeiro lugar gostaria de agradecer à Hikma Farmacêutica pela oportunidade de realizar o meu
estágio nas suas instalações.
Ao Samuel Camocho, orientador externo do estágio, por me ter acolhido e recibo da melhor forma
dando-me a excelente oportunidade de fazer parte da sua equipa, pelos seus conselhos e pela sua
disponibilidade para me ensinar.
Ao Professor Carlos Henriques, orientador interno da tese pela disponibilidade e orientação prestados
nesta fase final.
Não posso deixar de agradecer a todos os meus colegas de trabalho e supervisoras que desde o
primeiro dia estiveram sempre disponíveis para o esclarecimento de dúvidas. Aos que fizeram parte
do meu dia-a-dia na Hikma tornando-o mais fácil.
À minha família. Aos meus pais por serem inalcançáveis comigo, por todos os esforços feitos por
mim, por todos os ensinamentos, valores, educação, amor e incentivo transmitidos, por serem os
meus maiores pilares. Às minhas irmãs por serem as melhores do mundo, por me colocarem à frente
delas, por todo o apoio, amor e carinho. Sem o vosso apoio e presença na minha vida nada disto era
possível. Ao resto da família por sempre terem acreditado em mim e por me depositarem confiança.
Às minhas amigas de sempre pelos momentos partilhados ao longo deste percurso, por todo o apoio
e acima de tudo por estarem sempre presentes nos bons e sobretudo nos maus momentos da minha
vida, sem o amor transmitido por elas seria mais difícil.
Ao João, por teres sido a maior presença e apoio nesta etapa, nos momentos mais difíceis e nas
horas de maior fadiga, por toda a compreensão e paciência. Por todo o amor, dedicação e pelos
momentos partilhados.
Obrigada!
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Table of Contents
Preface.................................................................................................................................................... iv
Abstract .................................................................................................................................................. vi
Resumo ................................................................................................................................................ viii
Acknowledgments .................................................................................................................................. x
Table of Contents ................................................................................................................................. xii
List of figures ....................................................................................................................................... xiv
List of Tables......................................................................................................................................... xv
List of Acronyms ................................................................................................................................ xvii
1. Introduction ................................................................................................................................... 20
1.1. Hikma Farmacêutica S.A. .................................................................................................... 20
1.2. Regulatory Authorities .......................................................................................................... 21
1.2.1. Documentation ........................................................................................................ 21
2. Lyophilisation ............................................................................................................................... 22
2.1. Desired Characteristics of lyophilised products: .................................................................. 23
2.2. Advantages and disadvantages of lyophilisation process ................................................... 27
3. Process Validation ........................................................................................................................ 29
3.1. Different Methods of Validation ............................................................................................ 29
3.2. Process Validation Phases ................................................................................................... 31
3.3. Qualifications Steps ............................................................................................................. 33
4. Process Validation, Quality System and Quality Risk Management (QRM) ........................... 35
4.1. Risk Assessment .................................................................................................................. 35
4.1.1. Risk Assessment Methodology ............................................................................... 36
5. Technology Transfer..................................................................................................................... 39
5.1. Scale-up ............................................................................................................................... 39
6. Manufacturing Process of Lyophilised Products .................................................................... 41
6.1. Manufacturing Process ........................................................................................................ 41
6.2. Description of the Line and Facilities ................................................................................... 43
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7. Related Work: Improvement of Lyophilised Pharmaceutical Form ........................................ 46
7.1. Description of Product Y 5 g/vial and 10 g/vial .................................................................... 46
7.2. Problem Reported ................................................................................................................ 48
7.3. Life Cycle of Product Y for Injection, USP 5 g/vial and 10 g/vial ......................................... 50
7.4. Practical Case ...................................................................................................................... 58
7.5. Line Transfer and Scale-up for Product 5 g/vial – a Process Validation .............................. 61
7.5.1. Process Evaluation Activities and Results .............................................................. 62
7.6. Launching ............................................................................................................................. 67
8. Conclusion .................................................................................................................................... 68
8.1. Future work .......................................................................................................................... 70
9. References .................................................................................................................................... 71
10. Appendix ....................................................................................................................................... 74
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List of figures
Figure 1 - Steps Involved in Lyophilisation (Lyophilisation Cycle).
Figure 2 - Lyophilisation Process Cycle.
Figure 3 - Phase Diagram of Water.
Figure 4 - Design of a Lyophilizer.
Figure 5 - The Different Phases of Process Validation.
Figure 6 - Classification of the Risk Level.
Figure 7 - Evaluation of the Risk Priority.
Figure 8 - General Steps Involved in the Manufacture of Lyophilised Products.
Figure 9 - Lyophilised Manufacturing Process and Facilities.
Figure 10 - General View of the Filling Machine.
Figure 11 - General GLT Layout.
Figure 12 - General Load/Unload Line Description.
Figure 13 - Freeze-dried Product Y bulk solution at -4.0 ºC.
Figure 14 - Freeze-dried Product Y bulk solution at -0.7 ºC.
Figure 15 - Freeze-dried Product Y bulk solution at -0.3 ºC.
Figure 16 - Typical Bottomless Breakage of the Product Y 5 g/vial Presentation During
Lyophilisation.
Figure 17 - Life Cycle of Product Y for Injection, USP 5 g/vial and 10 g/vial.
Figure 18 - Squeme of Loading positions for PV batches and samples collection.
Figure 19 - Life Cycle of Product Y for Injection after Launching.
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List of Tables
Table 1 - Parameters for Production of Product Y.
Table 2 - First Submission Lyophilisation Cycle Parameters.
Table 3 - First R&D Trial Lyophilisation Cycle Parameters.
Table 4 - Second R&D Trial Lyophilisation Cycle Parameters.
Table 5 - Fourth R&D Trial Lyophilisation Cycle Parameters.
Table 6 - Final Lyophilisation Cycle Parameters to be Applied for FDA Submission.
Table 7 - Approved Freezing Phase Parameters.
Table 8 - Lyophilisation Cycle Parameters Comparison for Product Y 5 g/vial (Current and
Improved Freezing Phase) and 10 g/vial.
Table 9 - Side-by-side Inspection Results of Batch of Product Y for Injection USP 5 g/vial.
Table 10 - Percentage of cracked vials reject after Scale-up and Line Transfer.
Table 11 - Launching for Product Y for Injection USP.
Table 12 - Approved Batch Sizes for Product Y for Injection, USP 5 g/vial.
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List of Acronyms
ANDA
API
cGMP
CMAs
CPPs
CQAs
DQ
EMA
FDA
FLC
FMEA
FMECA
GMP
HEPA
INFARMED
IPC
IQ
MENA
OQ
PAT
PPQ
PQ
PV
QRM
R&D
Abbreviated New Drug Application
Active Pharmaceutical Ingredient
Current Good Manufacturing Process
Critical Material Attributes
Critical Process Parameters
Critical Quality Attributes
Design Qualification
European Medicines Agency
Food and Drugs Administration
Filling Machine BOSCH
Failure Modes Effect Analysis Method
Failure Mode Effect Critical Analysis
Good Manufacturing Process
High Efficiency Particulate Arrestance
Autoridade Nacional do Medicamento e Produtos de Saúde
In Process Control
Installation Qualification
Middle East and North Africa region
Operational Qualification
Process Analytical Technology
Process Performance Qualification
Performance Qualification
Process Validation
Quality risk management
Research and Development
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RPN
SA
USP
WFI
Risk Priority
Active Substance
United States Pharmacopoeia
Water for Injection
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1. Introduction
This dissertation under the theme, " Optimization of a Manufacturing Process of Parenteral Lyophilised
Drugs: Process Validation", was designed to obtain a master's degree under the Master of
Pharmaceutical Engineering, in which for six months, I did a practical internship at Hikma
Farmacêutica S.A.
I was part of the production team of lyophilised injectable products where I had the pleasure to follow
all production processes, documentation for each batch, people management, process optimization
and the opportunity to support the commissioning and validation of a new manufacturing line
production.
The purpose of this thesis is to contribute to the knowledge about development of injectable
pharmaceutical products and the improve/optimization of the process and how technology transfer and
Process Validation is performed at Hikma Farmacêutica.
1.1. Hikma Farmacêutica S.A.
In 1978, Hikma company was founded in Jordan by Samih Darwazah where it has established itself as
a leading supplier of branded generics and in-licensed products. After two years, on the 80’s, Hikma
established the first FDA-inspected manufacturing plant in the region of MENA (Middle East and North
Africa region). In this decade, Hikma extended beyond the Middle East and acquired land in Portugal
for the construction of a sterile manufacturing plant for injectable pharmaceutical products. The land
acquired in Sintra, Terrugem was the first step into a new region and major turning point for the
company.
After the successful expansion in Portugal and the continued effective delivering medicines in the
MENA, Hikma entity expressed interest in the US market and acquired West-Ward Pharmaceuticals,
in 1990s. The continuous growth of the company came with the acquisition in Tunisia and Saudi
Arabia who rename Hikma as a leading provider of generics in the MENA region but also
internationally.
In 2000, Portugal and Saudi Arabia manufacturing plants had successfully passed FDA inspections. It
was in this decade that Hikma expanded into the lyophilised market - variant of injectable products-
with the investment of a specialized manufacturing plant in Italy. Not yet satisfied, they invested on the
oncology market by buying Ribosepharm GmbH and Thymoorgan in Germany.
Hikma continued to expand, not just in Europe, but also in Egyptian market with Alkan Pharma and
Jordan with acquisition of Arab Pharmaceutical Manufacturing Company.
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After all the investments, 2005 was a historic mark and the Group listed on the London Stock
Exchange raising gross proceeds of US$124 million.
The current decade has seen growth through acquisitions of penicillin manufacturing plant in Al Dar Al
Arabia in Algeria.
Hikma saw a future on the injectables business market and bought Baxter’s, Bedford Laboratories and
Ben Venue Laboratories in the United States also as Roxane Laboratories in Columbus who helped
create a good position in the non-injectables market.
Nowadays, Hikma produces over 650 products and is present in 50 countries with 29 manufacturing
plants and seven different R&D centers.
In Portugal, the Group have three different facilities for parenteral drugs: Production of powders,
liquids and injectables and one recently implemented for the production of oncological products. Being
Sintra site known as the “center” of the injectable expertise for the company.
1.2. Regulatory Authorities
The pharmaceutical industry, through current Good Manufacturing Practices (cGMPʹs), maintains high
quality assurance standards with regard to the development, manufacture and control of medicines.
GMP also ensures that all products are manufactured only by authorized manufacturers whose
activities are regularly inspected by the competent authorities using quality risk management
principles.
Furthermore, Regulatory Agencies are responsible for controlling all activities concerning the
development, manufacture and marketing of pharmaceutical products. Given the fact that Hikma
Pharmaceuticals' largest target market is the United States, it is important to cite the Food and Drugs
Administration (FDA) regulatory agency. For this company, not only FDA is important but also the
agencies with national and European jurisdiction, INFARMED I.P. and European Medicines Agency
(EMA).
All European Community pharmaceutical manufacturers require manufacturing authorizations whether
the products are sold inside or outside the Community.
1.2.1. Documentation
Hikma is committed to high standards of ethical conduct and is required by law to create and maintain
a documentation system that involves all specifications, formulations, processing and packaging
instructions, procedures and records of the various manufacturing operations performed. Documents
must be clear, error free and up to date (INFARMED).
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2. Lyophilisation
Lyophilisation, also known as freeze-drying, is a very soft drying process widely used in
pharmaceutical industry to produce various pharmaceutical products including: parenteral products,
vaccines, proteins, antibiotics, serums and biotechnological products (Hottot, Vessot, and Andrieu
2007) (Abdelwahed et al. 2006).
Lyophilisation can be defined as the drying process of a frozen product in which the majority of water
and other solvents is directly removed by sublimation (converted directly into water vapor) without
passing through the liquid state leaving a dry porous mass of approximately the same size and shape
as the original frozen mass (Deluca and Lachman 1965; Nireesha et al. 2013).
Many pharmaceutical products as parenteral and biopharmaceutical products are thermal sensitive
(thermolabile) or unstable in solution form for prolonged storage periods, but that are stable in the dry
state (S. M. Patel, Doen, and Pikal 2010). Lyophilisation process is used in this products that might
lose their quality or activity in conventional evaporative drying, removing the solvent to promoting long-
term stability and increases the shelf life of the pharmaceutical drugs (Taylor et al. 2007; Nireesha et
al. 2013). Freeze-dried formulations not only have the advantage of better stability, but also provide
easy handling (shipping and storage) (Franks 1998; Heller, Carpenter, and Randolph 1997).
Freeze-Drying cycle can be divided into three steps: freezing (solidification), primary drying (ice
sublimation) and secondary drying (desorption of unfrozen water).
Figure 1. Steps involved in lyophilisation (Lyophilisation cycle).
Controlled lyophilisation keeps the product temperature low enough during the process to avoid
changes in the dried product appearance and characteristics (Ellab Validation Solutions 2018).
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2.1. Desired Characteristics of lyophilised
products:
• Intact cake;
• Uniform color;
• Sterile;
• Sufficient strength;
• Rapid dissolution (sufficiently porous);
• Chemically stable;
• Free of particles;
• Dried.
At the end of the lyophilisation a good appearance of dried cake with high quality and a porous
structure allowing a fast reconstitution after rehydration is desired.
Freezing stage
In the past, process optimization was mainly focused on the primary and secondary drying step.
However, with improved process understanding, it was recognized that the freezing step is also
important and start to be highlighted.
Freezing is the first step of lyophilisation. This process consists on converting the liquid solution into
the solid state in which water is converted to ice form by applying low temperatures. As the freezing
process continues, more water contained in the liquid freezes and the concentration of the remaining
liquid increases (called the freeze concentrate) (Tang and Pikal 2004).
The freezing phase is a critical step because it fixes the ice crystals structure, shape and dimensions.
The type of ice crystal structure affects the pores specific area, a key factor for the drying process and
rehydration time and consequently the sublimation time which is the longest time of the whole
lyophilisation process. The ice crystal sizes depend also on the vial size and type and also on the
filling height, larger fill volume takes longer to fully freeze (Hottot, Vessot, and Andrieu 2007;
Abdelwahed et al. 2006).
Some products are simple crystalline material, although the majority of products that are lyophilized
are amorphous and form glassy states when frozen. Frozen products can be categorized as crystalline
or amorphous glass in structure: amorphous solids are rigid structures but they lack a well-defined
shape, they are non-crystalline.
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Crystalline products have a well-defined eutectic freezing and melting point that is its collapse
temperature. Amorphous products have a corresponding glass transition temperature and they more
difficult to freeze dry. The collapse temperature of amorphous products is typically a few degrees
warmer than its glass transition temperature.
The determination of the critical collapse temperature of a product is an important step in establishing
and optimizing a freeze-drying process. This critical temperature determines the maximum
temperature that the product can withstand during primary drying without it melting or collapsing.
To optimize the lyophilisation cycle, optimal ice crystal sizes must be searched to minimize the
operating costs related to the whole duration of these drying steps. Usually, rapid cooling results in a
product with small ice crystals and more difficult to freeze-dry while slower cooling results in large ice
crystals.
The cooling rate and the supercooling degree are factors who control the freezing process and are
correlated with the morphology of the frozen material affecting the drying process. The supercooling
degree is defined as the difference between the equilibrium freezing point and the temperature at
which ice crystals first form. The supercooling degree is important because determines the number of
ice crystal formed (Constantino and Pikal 2004).
Freezing is a key step because microstructure established during this process usually represents the
microstructure of the dried product (Nireesha et al. 2013) and also has shown impact in intra-vial and
inter-vial uniformity as primary and secondary drying performance (Kasper, Winter, and Friess 2013).
Normally, vial breakage can be prevented by using slow freezing.
The cooling rate
The frozen liquid should be kept at the set temperature for sufficient time to transform all the
suspension into solid. Different ramped cooling on the shelves give different supercooling effects. One
practical approach to improve a lyophilised product consists in minimizing the specific surface area of
ice by growing large ice crystals; this can be possible by controlling the rate of super cooling. Normally,
higher supercooling rate results in larger ice specific surface are forming smaller ice crystals. A high
cooling rate induces higher nucleation of the ice crystal making compact ice crystal while a moderate
cooling rate is better for lyophilised products (Tang and Pikal 2004; Hottot, Vessot, and Andrieu 2007).
Primary drying
After all water and solutes have been converted into a frozen matrix, conditions must be established to
start the primary drying stage. In this process, the bulk water can be removed from the frozen product
via sublimation. Typically, the primary drying stage consumes the largest cycle time of lyophilisation
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and the optimization of this step has a large economic impact (Tang and Pikal 2004; Adelaide et al.
2004).
Once the product is frozen, primary drying starts whenever the chamber pressure is reduced by
vacuum pump evacuation and the shelf temperature is raised to supply the heat removed by ice
sublimation. Heat is transferred by thermal conduction from the shelf through the vial and finally gets
into the product. Sublimation requires heat energy to drive the phase change process from solid to gas
and it is important to maximize the surface contact of the vial with the shelf (Abdelwahed et al. 2006).
During this process, the chamber pressure is well below the vapor pressure of ice and ice is migrated
from the product to the condenser and crystallization onto the cold plates in the condenser (Tang and
Pikal 2004). The rate of sublimation ice depends on the difference in vapor pressure of the product
compared to the vapor pressure of the collector because the molecules are transferred from the high-
pressure sample to the lower pressure area. The concentration gradient between dried face and
condenser is the driving force for removal of the bulk water. In the condenser, a refrigerated surface at
a temperature below the product removes the vaporized solvent evolved by the drying material from
the vacuum chamber by converting it back as ice (Deluca and Lachman 1965).
The vapor pressure increases with an increase in temperature and are directly correlated. For this
reason, is necessary that the product temperature to be warmer than the collector temperature.
However, it is difficult to find an equilibrium because the temperature must be also low enough to
prevent any melting of the frozen mass maintaining the integrity and sufficiently high to maximize the
drying process and complete in a reasonable cycle time (Nireesha et al. 2013; Gan et al. 2005) .
Primary drying is a slow process in which product must be kept below its critical collapse temperature,
as the product degradation may occur, to produce acceptable drug product (S. M. Patel, Doen, and
Pikal 2010).
In the end, solute must form the rigid structure to support its weight after ice removal.
Secondary drying
The last stage of lyophilisation is the secondary drying. Secondary drying involves the removal of
adsorbed water from the cake which did not separate out as ice during the freezing step therefore did
not sublimate off.
At the end of primary drying, when all of the free ice crystals have been sublimed, the product will
appear to be dried. However, the moisture content can still have a percentage of water molecules
attached to the product. Usually, residual water is present in enough quantities to cause rapid
decomposition of the product when is stored at room temperature. Accordingly, is necessary to do a
secondary drying (Tang and Pikal 2004).
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When the secondary drying starts, the product and shelf temperature will increase and must be kept at
high temperature for a period sufficient to allow the isothermal desorption. Normally, for an
optimization of the process is better to run high shelf temperature for a short time than a low
temperature for a long period. Contrary to primary drying which use low shelf temperature and high
vacuum, secondary drying raises the shelf temperature (higher than ambient) and reduces the
chamber pressure to the lowest attainable level.
The residual water in the product remaining at this stage is stronger, requiring more energy to be
removed.
When lyophilisation is over, the moistures contents should be less than 5%, resulting in an elegant
porously dried cake (S. M. Patel, Doen, and Pikal 2010).
Figure 2. Lyophilisation process cycle.
The Principle of Freeze-Drying
At the triple point liquid, solid and gas coexist in equilibrium. When the water is heated below the triple
point, then it will change from solid state to vapor state directly. Sublimation can take place at
pressures and temperature below triple point to enable conversion of ice into vapor, without entering
the liquid phase.
The eutectic temperature is the temperature at which a product start to melt into a liquid. During
lyophilisation, a product must be held below the eutectic temperature for sublimation to occur from the
solid state. The first step for lyophilisation is freezing the product bellow the eutectic temperature.
Then in primary drying phase the temperature raises near to the collapse temperature with a safety
margin and is subjected to a reduced pressure turning the water content into vapor leaving solid and
dried components of the original liquid (Nail et al., 2002).
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Figure 3. Phase Diagram of Water.
2.2. Advantages and disadvantages of
lyophilisation process
Advantages:
• Ensures sample stability in a dry state;
• Removal of water without excessive heating of the product
• Rapid and easy dissolution of reconstituted product (rapid reconstitution time);
• Increases shelf life;
• Easily for storage (can be stored at room temperature) and shipping (reduces weight and volume of samples);
• Good for O2 and air sensitive drugs;
• Product is process in the liquid form (simplifies aseptic handling);
• Does not shrink samples;
• Purity of samples
• Elegant cake appearance
Disadvantages:
• Increased handling and processing time;
• Cost (expensive unit operation) and complexity of equipment;
• Issues associated with sterilization and sterility assurance of the dryer chamber and aseptic loading of vials into the chamber (Drug Discovery and Development);
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Figure 4. Design of a lyophilizer
The lyophilisation process occurs in a lyophilizer or freeze-dryer (figure 4). The main components are:
• Chamber;
• Condenser;
• Vacuum pump.
A lyophilizer consists of a chamber that contains product shelves capable of cooling and heating the
vials. A vacuum pump, a condenser, a refrigeration unit and associated controls are connected to the
vacuum chamber.
The most common application of lyophilisation is the manufacturing of parenteral products that are
administered after a simple reconstitution step (Shukla 2011).
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3. Process Validation
Effective process validation contributes to assuring drug quality (FDA, 2011). The basic principle of
process validation “is the collection and evaluation of data, from the process design stage through
commercial production, which establishes scientific evidence that a process is capable of consistently
and uniformly producing a product according to the specifications and quality characteristics” (FDA,
2011).
Moreover, process validation assures that product quality is performed, within established parameters,
consistently from batch-to-batch and unit-to-unit (WHO 1996). The purpose of validation is to prove the
effectiveness in a manufacturing process to ensure varied inputs with high quality outputs (V. K. K. B.
Patel and Pethe 2010; ICH Definition).
A well-established PV results in a reduction of product losses, incidence of deviations, quality cost,
down times and the risks of failing in GMP. Additionally, increases the safety levels, the data to provide
knowledge for a robust process, greater rationalization of the activities developed, simpler
maintenance of equipment and easier scale-up from development work (Nandhakumar 2013).
3.1. Different Methods of Validation
Currently, validation is conducted according to different approaches: based on experimental data or
historical data.
Based on experimental data we have prospective validation and concurrent validation. The first one
happens before process commissioning or the product's market reach, while the latter is executed
during production (Comissão Europeia 2001). Retrospective validation is centered in historical data
and used for established products with stable processes.
Prospective Validation
Prospective validation is adopted when new drug products are introduced, a product is made under a
revised manufacturing process or before the process is launch in commercial use. A risk analysis of
the production process is made during the development stage and is examined into individual steps to
determine if they might lead to critical conditions that may affect the quality of the product. This
evaluation is supported by data from experimentation and theoretical considerations.
Three different experiments have to be designed and entirely reported in a protocol. Equipment,
production environment and analytical assay methods need to be validated and critical situations
identified. To have a PV well designed, an extensive sampling and testing should be done at different
steps of the manufacturing process.
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In case unstable conditions are discovered the risk must be evaluated and the potential causes are
considered. Inadequate manufacturing processes are modified and improved until validation proves
that they are acceptable and finally master batch records can be made attending all the critical
conditions and parameters determined (Ahir et al. 2014; Nandhakumar 2013).
Concurrent Validation
This validation occurs during normal production runs and involves in-process monitoring. Concurrent
validation supports documentation to prove that the production process in under control. The decision
to perform this validation needs to be correctly justified, well-reported and approved by authorized
labourers.
Retrospective Validation
is based on historical data, documents that provide evidence for a structure's/system's correct
operation and control. This type of validation is established for products with stable processes and
already in distribution. If the operating procedures, the composition of the product or the equipment
are changed or reviewed this type of validation shouldn't be applied (Ahir et al. 2014; Nandhakumar
2013).
The required protocol with the results of the data review should include batch processing and
packaging records, maintenance logbooks, process control charts, records of personnel changes,
finished product data, storage stability results and process capability studies (WHO 1996).
Retrospective validation is no long encouraged and prospective validation is opted for. In fact, the
impossibility to use retrospective validation in manufacturing of sterile products has a negative impact
which leads to preferences in other types of validation.
Revalidation
Revalidation, used after a change with impact on product quality, serves to demonstrate that any
intentional or unintentional change in a process manufacturing or in the process environment does not
adversely perturb process parameters and quality characteristics. Documentation requests are the
same as for the initial validation of the process.
After a change, it is important to implement the alterations affecting the standard procedure to confirm
they maintain validated and high quality. Whoever facilities, equipment and manufacturing techniques
including cleaning where no significant modifications have been made, needs a periodically review to
confirm they remain valid. Each alteration must be carefully analysed by validation department to
determine if it is sufficient to opt for a revalidation and in that case, in which scope (Ahir et al. 2014;
Nandhakumar 2013).
Some situations that require revalidation are:
• Changes in raw materials (that can affect the process or the product quality);
31
• Changes in the manufacturing process- different batch size;
• Changes in the equipment;
• Changes in the facilities;
• Changes in the provider of the active raw material manufacturer;
• Changes in packaging material.
In cases of replacement of the equipment for exactly the same machine normally is not require
revalidation except the new equipment need to be qualified for the use.
A well justified document is necessary to support the decision of not perform revalidation studies and
must be done by authorized persons.
3.2. Process Validation Phases
Validation in general requires a meticulous preparation and careful planning involving various activities
during the lifecycle of the product and process. PV is divided in three phases: process design,
process qualification and continued process verification.
Figure 5. The different phases of Process Validation.
Process Design
PV is described as the collection and evaluation of data. In this phase, is exclusively focused on
qualification efforts and the manufacturing process is drawn based on knowledge gained during R&D
and scale-up activities.
During the research and development phase, important aspects as formulation, dosage forms, stability
and storage conditions, process capability, equipment qualification, pilot batch, scale-up and transfer
technology studies contribute to the process design (Ahir et al. 2014; Nandhakumar 2013).
Process control strategy takes place in this phase.
32
Process Qualification
Process Qualification is where process design is evaluated to check if the process is capable to
reproduce commercial manufacturing batches. There are two different aspects of process qualification
to be considered:
• Design of the facility and qualification of the equipment and utilities;
In this stage, activities are performed to demonstrate that all equipment and utility systems are
suitable for their intended use and to verify if are capable to perform properly. The first step of these
activities is the selection of the equipment and utilities, the construction materials, the operating
principles and the performance characteristics.
The second step is the Installation Qualification (IQ). The IQ exists to verify if all equipment is built and
correctly installed in compliance with the design specifications, if have all capacity and functions
requested and if it is correctly calibrated and connected.
Operational Qualification (OQ) follows IQ and is used to confirm that utilities and equipment operate in
accordance with the specifications and operating ranges.
• Process Performance Qualification (PPQ).
In this stage, GMP procedures should be followed. It is essential evaluate if the process design is able
to reproduce commercial manufacturing batches. After properly tested, even under “worst-case”
conditions, with satisfactory results and all established limits of Critical Process Parameters are
validated it is possible to start the commercial distribution (Ahir et al. 2014; Nandhakumar 2013).
Continued Process Verification
On-going verification it is required during production routine to confirm that the process remains in a
stable and solid control.
Data collected during continued process validation is important to optimize the process or to improve
the quality of the drug. When corrective actions are made due to problems are detected during this
phase, a document describing the changed plan must be written with a rationale for the adjustment, a
good strategy implementation of the change plan and it has to be approved by the quality department.
To assure continued process validation a frequent review of all process related documents including
failures, deviations, change control procedures and process modifications as also validation audit
reports are studied (Ahir et al. 2014; Nandhakumar 2013).
33
3.3. Qualifications Steps
For a good work in a pharmaceutical industry it is imperative for the qualification to keep up with
validation. Validation is the “act of prove effectiveness” in a process while qualification is used to
evaluate and qualify an equipment. A process is considered validated when the equipment and
process are able without put in risk the safety and quality of the product.
For this reason, validation and qualification must work synchronized and in harmony (WHO 2006,
Sharma 2013).
Design Qualification (DQ)
In new facilities or equipments, the first section of validation is the design qualification. The design
must be established in compliance with GMP and well reported (Saudi Food & Drug Authority 2010).
Installation Qualification (IQ)
The main goal of this type of qualification is to check if all processing equipments are in accordance
with the installation specifications (manuals and engineering drawings). IQ is executed on new
facilities, systems and equipments and must include the examination of equipment design, the
maintenance adjustments, the equipment calibration, the services and instrumentation checked, the
working instructions, the verification of materials construction, the description of equipment, the
principle of operation and equipment requirements.
In short, IQ is to demonstrate that the installation was successfully completed.
Operational Qualification (OQ)
Operation qualification follows a previously authorized protocol. In this document, OQ protocol should
identify the studies to be undertaken on the critical operating conditions, the acceptance criteria to be
met, the sequence of those studies and the measuring equipment to be used. OQ should take in
account the circumstances, covering upper and lower processing and the upper and lower limits
mentioned as “worst-case” scenario including operation controls and alarms. OQ should include tests
that challenge the equipment and it’s functions and also incorporates the stoppages, interventions, the
start-up and all activities expected during the routine production.
34
The main goal of OQ qualification is to prove satisfactory operation over the normal operation routine
procedure and consequently acknowledge the basis for training the operators (Ahir et al. 2014;
Nandhakumar 2013).
Performance Qualification (PQ)
After a successful IQ and OQ, the final step (performance qualification) should be complete to certify
that the equipment is working reproducibly and safety within a certain set of parameters for the critical
variables.
Tests using production materials and tests using a set of conditions to simulate worst-cases, cleaning,
calibration and preventive maintenance need to be reported.
In order to produce PQ batches (normally corresponding to the first commercial-scale batches of the
drug product) a control strategy for the manufacturing process should have been established and
previously well-designed.. It is important to define the performance criteria, how much and what data
must be collect and when data must be collected
Usually, a PQ protocol has a higher level of sampling, additional testing of process manufacturing than
during routine commercial production. The increased level of examination and sampling is used to
establish appropriate levels and frequency for the routine sampling and monitoring, taking in
consideration the process complexity, the size of the batch, the previously experience with the product
and process.
After a successful PQ comes the commercial distribution of the drug product. All data gathered from
commercial-scale batches and also from laboratory and pilot-scale studies is used to support the
decision of beginning commercial distribution (Ahir et al. 2014; Nandhakumar 2013; WHO 1996).
35
4. Process Validation, Quality System
and Quality Risk Management (QRM)
Before a batch is commercialized, the manufacturer must demonstrate that the commercial
manufacturing process is capable of consistently produce acceptable quality products within
commercial manufacturing conditions - meeting those attributes relating to identity, strength, quality,
purity, and potency (CQAs). The assurance should be obtained from objective information and data
from laboratory, pilot, and/or commercial scale studies. (Guidance for Industry – Process Validation:
General Principles and Practices, January 2011).
Process validation is part of technology transfer and is used to demonstrate that the manufacturing
process developed by R&D and transferred to industrial scale, operated within established
parameters, can consistently yield a product meeting its specified quality attributes. It is important
however to recognize that process validation is not an isolated event.
The purpose of process validation is to certify a set of varied inputs cannot affect the regular
production and lead to consistent and high-quality yields. PV is an ongoing process throughout routine
production that must be adapted by pharmaceutical companies as a guaranteed quality and safety
manufacturing control.
Quality risk management is a systematic process that identifies and prioritizes, evaluates and controls
potential risks to quality. It facilitates continual improvement (assessment, control, communication and
review of risks) of process performance and product quality throughout the product lifecycle.
Risks identification is a key step for the safe design of a manufacturing process; after the threats are
pointed out it is important to evaluate their consequences, as well as their causes.
4.1. Risk Assessment
The main goal of risk assessment is to minimize the process risk based on the identification of the
potential risk sources. The risk assessment has to be part of a quality risk management system and
ask the following questions:
- What can fail?
- What is the probability that the failure will occur?
- What are the consequences if this process goes wrong?
The first phase of risk assessment is the risk identification and consists in answering the question:
“What can cause the problems?”. The goal is to make a list with all potential risks and can be
extensive as possible. Since identifying all potential failures can be an enormous task when
36
considering an entire manufacturing process, the process map generated during the initiation phase of
the risk management process is a tool to do logical and focused steps during the risk assessment.
After the risk identification starts the risk analysis and involves a risk assessment tool , which have a
qualitative or quantitative estimation of the consequence and the ability to detect the failure.
Next the risk analysis, a risk evaluation is required to measure if the risk is in an acceptable level of
risk (the action threshold) or not. At this point, the risk assessment phase ends. The risk management
process continues through the steps of risk control, risk review and communication. This implies that
during lifecycle of the product, the risk assessment is reviewed. This review includes adjustments to
regulatory requirements or to additional information related to the new process.
4.1.1. Risk Assessment Methodology
The risk assessment is made on the Failure Modes Effect Analysis Method (FMEA). This method is
based on the identification of the potential risk sources – defined as Failure Mode. Each failure mode
is evaluated and classified as result of the Probability of Occurrence [P], Severity of the Potential effect
of the failure [S] and the Detectability [D]. The risk prioritization is calculated as RPN = [P] X [S] X [D].
Frequency of probability [P]
The probability of an event may be rated, but not necessarily limited to, 1, 2, or 3 according to its
likelihood.
• Level 1 (low): The frequency of the event occurring is perceived low;
• Level 2 (Medium): The frequency of the event occurring is perceived to be medium;
• Level 3 (High): The frequency of the event is perceived to be high.
Severity of the potential effect of the failure [S]
This approach requires the team to consider which impact this change/event has on the product
quality. The impact of the consequence may be rated but not limited to1, 2, or 3:
• Level 1 (Low): Expected to have a minor negative impact on the product quality.
• Level 2 (Medium): Expected to have a moderate impact on the product quality.
• Level 3 (High): Expected to have a very high significant negative impact the product quality.
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Probability of detection [D]
The purpose of this stage is to identify if the risk can be recognized or detected by other means in the
system. The probability of a risk being detected is rated 1, 2 or 3 according to its detection
possibilities:
• Level 1: reliable detection methods are in place (validated methods of analysis).
• Level 2: there is a reliable detection used but are not designed to properly detect the quality of
the product parameter that is being tested.
• Level 3: there is no reliable detection method available to test the product quality parameter in
question.
Risk Classification [C]
Having assigned the probability of the risk and the level of impact that such an event may have, the
risk can be classified. The risk is a multiplication of probability [P] and severity [S]. Therefore, the risk
ranges from:
Figure 6. Classification of the risk level.
The Risk Classification (PxS=C) gives us an indication of the impact on product quality or data
integrity:
• Level 1 means that the probability that this failure appearing is high and that the impact on
product quality is high or medium;
• Level 2 means a moderate impact on product quality.
• Level 3 means that we have practically no impact on product quality.
Risk Priority [RPN]
The risk priority is a multiplication of three parameters, thus taking in account the potential failure [C;
Risk Classification] associated with the potential effect and its detectability [D]. The risk priority can
range from:
38
Figure 7. Evaluation of the risk priority.
The Risk Priority is used to select the appropriate risk mitigation and to focus the validation effort:
• A high priority RPN means that this function or component is critical and other measures and
controls need to be taken to mitigate the risk.
• A medium priority RPN means that this function or component is potentially critical and other
measures and controls needs to be in place to mitigate the risk.
• A low priority RPN means that this function or component is non-critical and that there are no
actions need to be taken.
Design of freeze-drying process is often approached with trial and error experimental plan or the
protocol used on the first laboratory run is adopted without further attempts at optimization, losing
robustness and efficiency (Tang and Pikal 2004).
All changes that may affect product quality or reproducibility of the process should be formally
requested, documented and accepted. The likely impact of the change of facilities, systems and
equipment on the product should be evaluated, including risk analysis.
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5. Technology Transfer
Technology transfer implicates transfer of product and process knowledge to achieve the
manufacturing process for commercialization and it includes all the activities required since the
pharmaceutical development (R&D) to production, for new products, or from one manufacturing site to
another.
During the technology transfer, many aspects need to be taken into consideration to allow proper
evaluation and validation of the manufacturing process before starting routine production for
commercialization. A proper analysis of material attributes and process parameters, inputs,
manufacturing controls, the process outputs and their quality attributes are essential for successful
technology transfer and an inappropriate understanding of this correlation may lead to an uncontrolled
process. An uncontrolled process can result in extensive product losses, batch rejection, difficulties
with regulatory submissions and lately in a non-approval submission.
Through a technology transfer, all unit operations and their associated equipment should be identified
as well as all the parameters and attributes considered critical. All relevant information about the
product and the process gained during the development phase, should also be properly reviewed and
evaluated.
When a lyophilised product is to be manufactured, an appropriate lyophilisation cycle should be
developed. During R&D the cycle development is done, however adjustments in cycle times and
parameters may occur when transferring from R&D freeze dryer to an industrial lyophiliser.
Lyophilisation involves many parameters (essential to achieve for a proper process control and to
obtain a cake with desired characteristics). These operation parameters include heat and mass
transfer, load condition (partial or full load), the container system, shelf temperature and chamber
pressure that are essential to achieve for a proper process control and to obtain a cake with desired
characteristics. These parameters sometimes differ with the technology transfer. Therefore,
manufacturing of test batches is advisable before production of the PPQ batches (Rogers, Takegami,
and Yin 2001).
5.1. Scale-up
Scale-up is generally defined as the process of increasing the batch size. Scale-up of a process can
also be viewed as a procedure for applying the same process to different outputs (volumes).
Scale-up of pharmaceutical manufacturing processes demand a combination of experience, science
and engineering. Therefore, for a successful scale-up is necessary the critical material attributes
(CMAs) and critical quality attributes of (CQAs) the products and the critical process parameters
(CPPs) of the equipment used in manufacturing process are well-defined.
40
Risk management tools can help controlling the critical steps of the process to achieve a successful
scale-up.
Three different scales exist: the laboratory scale batches, following the pilot batches scale and the
production batches scale.
Laboratory Scale Batches
The traditional process of pharmaceutical product development until it reaches the global market
involves a long journey of many experiments, observations, challenges, and resolutions. These
batches are produced at the research and development (R&D) laboratory stage and they present a
very small size.
During the initial formulation stage, certain parameters are considered to convert the raw materials
into a drug formulation with an aim to maintain the required quality attributes. However, at the initial
level, these investigations are performed at a small scale by using small-output equipment, where the
methods and results observed from these investigations are appropriate to that scale only (Amirkia
and Heinrich, 2015).
The production of R&D batches is helpful to support formulation and packaging development, clinical
and pre-clinical studies. The data that these batches provide are useful to define the products
characteristics and enable the choice of appropriate manufacturing process (Nandhakumar 2013).
Pilot Batches
Moving from R&D to production scale, it is sometimes essential to have an intermediate batch scale-
Pilot Batches which is defined as the manufacturing of a product by a procedure fully representative of
and simulating that for manufacturing scale.
These batches are helpful to gain knowledge and also make possible the production of enough
product for clinical testing and samples for marketing. However, inserting an intermediate step
between R&D and production scales does not in itself guarantee a smooth transition. A well-defined
process may produce a perfect product in laboratory and pilot scale and then fail quality assurance
tests during production.
Production Scale Batches
The batches are of the normal size which will be produced during the routine marketing of the product.
Data on production scale batches may not always be available prior to granting marketing
authorization. When production scale data are not available it is necessary the evaluation and
41
characterization of the critical process parameters at laboratory or pilot scale followed by the
manufacture of submission batches and to complete a validation program on production scale batches
(Levin, 2001).
6. Manufacturing Process of Lyophilised
Products
Parenteral drugs are preparations intended for injection through the external tissue, for example the
skin, so that the active substances are administered using gravity or force directly into a blood vessel,
organ, tissue or lesion. These drugs are available as liquid (solutions, emulsions or suspensions) or
solid products (powder – cephalosporins or lyophilised products). Injectable pharmaceutical products
are prepared by methods properly designed to ensure safety and quality and to meet Pharmacopeial
requirements for sterility, pyrogens, particles and contaminants.
6.1. Manufacturing Process
The manufacture of pharmaceutical products should follow the requirements of the current GMPs. The
adherence to cGMPs assure a proper design, monitoring and control each step of a process and the
correct facilities. The use of GMPs guarantees the quality, safety and purity of drug products by
requiring that is manufactured in a properly way and strictly controlled each operation process.
The manufacture of lyophilised products must occur according the sterile rules in clean areas,
constantly cleaned following a standard, and supplied with air who has passed through HEPA filters.
The sterile pharmaceutical products can by produced by terminal sterilization or aseptic processing.
The aseptic processing method is just acceptable when terminal sterilization is not possible due to the
degradation of the drug product when exposed to terminal sterilization conditions, as is the case with
lyophilised products.
Aseptic Processing
Sterile drug products can be manufactured using two techniques: terminal sterilization or aseptic
processing. Terminal sterilization usually involves heat or irradiation. However, some drugs can be
destroyed by exposure to heat or irradiation and it is required to use aseptic filling.
In aseptic filling the drug product and container/closure are subjected to sterilization methods
separately, as appropriate, and then brought together. Because there is no process to sterilize the
product in its final container, it is crucial the containers be filled and sealed in an extremely high-quality
42
environment. Before aseptic assembly into a final product the individual parts of the final product must
be subjected to various sterilization processes (Sandle, 2011).
All the components of the final product are sterilized. The rubber stoppers are subjugated to moist
heat sterilization in an autoclave or acquired irradiated (pre-sterilized), the glass containers pass
through a depyrogenation tunnel – dry heat sterilization- and the liquid product subjected to one or
more sterilization filtrations under Grade A. All these manufacturing processes are individually
validated and continuous controlled.
Figure 8. General Steps Involved in the Manufacture of Lyophilised Products.
The first step of the manufacturing process of lyophilised products is the dissolution of the active
substance ingredient (API) and the excipients in the water for injection (WFI) or another solvent. After
the compounding, the bulk solution must pass through a 0,22 µ bacterial retentive filter to sterilize the
product.
While the bulk solution is prepared, the vials are washed to remove the particles and submitted to dry
heat sterilization passing by through a depyrogenation tunnel. Each individually container is filled with
the sterile bulk solution and partially stoppering the containers under aseptic conditions (Grade A).
43
The partially stoppered vials filled will be transported to the lyophilizer and loading into the chamber
under aseptic conditions. After that, the solution first will be freeze by placing on cooled shelves in a
freeze-drying chamber and secondly vacuum will be applied to the chamber heating the shelves in
order to sublimate the water from the frozen state. When all the water is sublimed from the vials a
complete stoppering will be done by hydraulic or screw rod stoppering mechanisms installed in the
lyophilizers. Finally, the stoppered vials will be transported from the freeze-drying chamber to the
capper to seal.
Thus, a compound that is heat-sensitive and not sufficiently stable in aqueous solution can be
formulated into a stable solid form, rapidly soluble and elegant injectable preparation (Deluca and
Lachman 1965).
Figure 9. Lyophilised Manufacturing process and facilities.
6.2. Description of the Line and Facilities
To describe the manufacturing process, it is necessary to do an approach of the lyophilisation
manufacturing facilities for the filling of aseptic sterilized products in the Injectable Division at Hikma I.
The batch is prepared in compounding room using the preparation tanks. During and after the
preparation, samples are taken for in process control (IPC) testing. After the compounding, the product
is transferred from the preparation tank to the transfer tank and is connected to the filling machine
(FLC) through a sterilizing grade, integrity tested, filter.
The product is supplied to the filling system via intermediate filling tank. The filling is controlled via
valve, normally closed, and when the valve opens the intermediate filling tank is filled using the
overpressure in the on-site product tank.
The washer for the vials is coupled with the depyrogenation tunnel, in grade D, and is connected with
a turntable of the FLC, under grade A, were the vials after washed and submitted to dry heat
sterilization, wait to be filled.
Filler and Stopper
Pré-Filter
Autoclave
Washer +
Tunnel
Compounding
Vials
Parts
Stoppers
Frames
Grade C Grade D Grade A
Lyophilization
Grade D
Capper100 % Visual Inspection
Pharmaceutical
Area
Under Grade A
44
The intermediate tank is connected to six pumps and consequently to six needles who reach the fill
volume to the vials who are running in a transition belt.
The FLC have an integrated monitoring level to control the IPC. Some containers are removed from
the transport belt with the IPC starwheel and weighed in a platform then are placed back into the filling
station and filled. After the filling station, the same vials are taken back to the gross weighing station
and tared. The measure of the values is to check if the vials filled can properly continuous on the
route, in case the fill volume is not correct the containers are not stoppered and in a station latter they
will be rejected from the transition belt.
Figure 10. General View of the Filling Machine.
For the products sensitive to the oxygen, the gassing system can be used to reduce the residual air
inside of the container by flushing to the interior of the vials a protective gas. The protective gas is
supplied by an infrastructure from an on-site system.
The stopper lock is used to release a stopper to the stopper-placing wheel when a vial is present that
should receive a stopper. The vacuum is used to hold the stoppers in the stopper-placing wheel and
the wheels move synchronic with the vial main transport. The wheels of the stopper station are taken
the container from the main transport and partially setting the stopper on the container and take it back
to the main transport.
After the stopper station, the vials are inspected by two sensors, in order to detect the presence of the
stoppers and to detect if stoppers are too high. The reject station is used to reject bad containers: low
volume, high volume, vials without stopper or not correctly placed.
45
The good partially stoppered vials are supplied by an in feed belt to the GLT Vials Tray Loader where
are stacked into rows. Once the tray has received the number of the rows determined by the recipe,
the vials are pushed into a tray and thereafter placed to the Lyo trays loader machine (MOTUS).
Figure 11. General GLT layout.
The lyophilisation trays loader machine (Motus) is used for the loading of trays filled with partially
stoppered vials from GLT to the lyophiliser and after the lyophilisation process, to unload the trays to
the turntable in feed of the capping machine. During freeze drying, the containers are totally stoppered
and when all lyophilisation stages are finished the trays will be unloaded by Motus and placed to the in
feed zone of the capping machine.
Figure 12. General load/unload line description.
Stako system is used to separate the vials from the trays for the turntable in feed of the capping
machine, while is collecting and transferring the trays for storage. The stoppered vials are transported
on a conveyor belt to be capped, under grade A air supply. After all the containers are sealed, they
proceed to the visual inspection for defects, ultimately labelling and final packaging, in the packaging
area.
1.1.1.
Section 1 2 3 4 5
Machine Bosch Vials Tray
Loader
Motus Semi-
Automatic Loading System
Hull
Lyophilizer
Stako Vials/Frames
Separation Station
Genesis Vials
Capping Machine
46
7. Related Work: Improvement of
Lyophilised Pharmaceutical Form
Due to confidentiality with Hikma Farmaceutica, S.A., this work will not mention the name of the Active
Substance (SA), or the dosages or the commercial name of the finished product, referring only to the
pharmacotherapeutic group to which it belongs and the pharmaceutical form. The drug studied will be
called Product for Injection Y.
7.1. Description of Product Y 5 g/vial and 10 g/vial
Product Y Lyophilised powder for Injection is a new product developed by Research and Development
Department of Hikma Pharmaceuticals at Jordan where the R&D is performed.
Product Y is for intravenous injection, a parenteral, indicated for treatment of severe infections and it is
a generic transferred to Hikma Farmacêutica manufacturing facilities in Portugal. The product Y, is
almost white, or tan to brown, free flowing powder, odourless and having a bitter taste, hygroscopic
powder, freely soluble in water and insoluble in ether and in chloroform.
The formulation is a simple aqueous-based formula that includes the API as well as an anti-gelling
agent and water for injection (WFI), both disappearing during lyophilisation process. The bulk solution
concentration for both strengths is the same 166.67 mg/ml and the pH should be between 2.5 – 4.5.
Studies indicate that Product Y is sensitive to light, unstable under high humidity, unstable under high
temperature and sensitive to oxygen.
The reconstitution volume is 100 ml for the 5 g/vial presentation and 95 ml for the 10 g/vial
presentation using sterile WFI.
Product Target fill volume Batch size Vial size
Injection Y (5 g/vial) 30.50 mL 40 L 100 mL, neck 20 moulded
Injection Y (10 g/vial) 61 mL 80 L 100 mL, neck 20 moulded
Table 1. Parameters for the production of Product Y.
47
During the characterization of the Product Y it was used the Freeze-drying microscopy (FDM) a
technique for identifying critical formulation parameters and is the only method for reliably determining
collapse temperatures. Using the FDM, the sample is firstly frozen until it reaches a temperature of -
40ºC. Then, the vacuum pump is turned on and the drying begins. The temperature is then increased,
slowly to allow to see the differences in behavior of the analyzed product.
Figure 13: Freeze-dried Product Y bulk solution at -4.0 ºC. The dried part is beginning to suffer bigger
breakage, whilst the matrix is still holding its structure.
Figure 14: Freeze-dried Product Y bulk solution at -0.7 ºC. As get closer to 0ºC, solution suffers
complete collapse.
48
Figure 15: Freeze-dried Product Y bulk solution at -0.3 ºC. The matrix melts completely.
In crystalline solutions the Critical Temperature for Freeze Drying is the eutectic point and in
amorphous products this is the collapse temperature.
The instability of the frozen matrix begins at approximately -5ºC/-4ºC. Because no crystalline shape is
manifested, and no eutectic behavior present (solid and liquid phases presented simultaneously), the
Product Y is an amorphous structure.
7.2. Problem Reported
In January of 2019 after the approval of FDA, Hikma Farmacêutica started to produce Product Y for
Injection, USP 5 g/vial and 10 g/vial and, for launching the product in the US Market and consequently
in order to validate the scale-up process, three Process Validation (PV) batches of 170 L in 400 L
tanks of Product Y for Injection USP 5 g/vial were manufactured at Liquid & Lyophilised Injectable
Production Department- Line 1 at Hikma Farmacêutica Portugal.
To perform all process validation activities, it was necessary more two PV batches under the scope of
PV protocol. The PV included compounding of the bulk solution, pre-filtration through a 1.2 µm filter,
filtration through sterilizing 0.2 µm filter, filling, lyophilisation, stoppering, capping and 100% visual
inspection. In the end, a side by side comparison of finished product results was performed between
40 L batch and 170 L batches manufactured in Line 1 in order to evaluate the reproducibility of results
or if any manufacturing constraint related to the scale up process is noticed.
The evaluation of the effects of the lyophilisation on the quality of the filled vials, lyophilisation
uniformity and the evaluation of the results of 100% visual inspection of the vials was performed in
49
three batches. During capping, in one of the inspection spots in the capper, it was noted that a
significant percentage of cracked vials were rejected (between 13.6% and 8.8% of the unloaded
quantity). After the amount of rejected vials was verify it was necessary to open a variance notice and
the batch was immediately put into retention and therefore it was not possible to perform 100% visual
inspection for the PV batches.
The significant percentage of cracked vials rejected turned the manufacturing process in a non-robust
and irreproducible process becoming invalidated. For that reason, the improvement of Product Y 5
g/vial as well as the identification of the root cause for the broken vials was necessary.
During the investigation it was possible to confirm that the problem of vial breakage was in the
freezing phase. The pattern of the crack (Figure 13) indicates that it is caused by expansion of the
cake during freezing phase due to the fact that freezing occurs rapidly and in one shot shocking.
The fast-freezing makes the system of the matrix to expand suddenly without allowing the crystals, for
enough time, to be shaped according to the vial size creating pressure on the edges and causing that
bottomless breakage of the glass followed by a crack on the wall of the vial (Figure 13).
Large fill volumes are strongly correlated with higher percentage of vial cracks especially with fast
temperature gradient. The strain on the vial's axial direction is significantly higher than the hoop
direction typically resulting in bottom lens of the vial coming off (Jiang, 2007).
Figure 16. Typical Bottomless Breakage of the Product Y 5 g/vial Presentation during Lyophilisation.
To follow this investigation, it was necessary to review all data collected and analyze the life cycle
history of the product.
50
7.3. Life Cycle of Product Y for Injection, USP 5
g/vial and 10 g/vial
Figure 17. Life Cycle of Product Y for Injection, USP 5 g/vial and 10 g/vial.
In 2011, after the R&D team prepared the formulation and studied the manufacturing process Hikma
Farmacêutica decided to propose to the authorities a new product: Product Y for Injection USP 5 g/vial
and Product Y for Injection USP 10 g/vial. To implement this product in the installations was defined
activities intended to evaluate the manufacturing process for this product for US Market.
A Risk Analysis and Process Validation studies were made to provide evidence that a defined
manufacturing procedure will consistently yield product meeting its predetermined specifications and
quality characteristics (Appendix 1).
Based on Process Validation, one submission batch of 40L was manufactured for Product Y for
Injection, USP 5 g/vial and one batch of 80L for Product Y for Injection, USP 10 g/vial. The submission
batches of Product Y for Injection USP 5 g/vial (40 L) and Product Y for Injection USP 10 g/vial (80 L)
were prepared on 23rd October 2011 and 24th October 2011 respectively at Hikma Farmacêutica
Portugal.
During the primary drying stage in submission batch for Product Y for Injection USP 10 g/vial (80 L),
the product was visually inspected through the sight glass located in both sides of the freeze dryer and
signals of breakage were observed. However, as the cycle was continued and, after the unloading of
the lyophiliser, around 90% of the obtained vials were broken.
In order to completely overcome the problem of vials breakage, R&D department in Hikma
Pharmaceuticals-Jordan started to investigate and to improve the problem appeared in the submission
batch. This research included different optimized cycles (additional trials) performed at R&D level and
industrial scale. After all changes, when R&D create a robust cycle with no vial breakage a new
submission batch of Product Y for Injection, USP 10 g/vial was applied.
The reason behind vial breakage was attributed to the fast-freezing rate applied during freezing phase
on the lyophilisation cycle. To support that, two 20 L batches (Trial #1 and Trial #2) were produced by
applying new optimized lyophilisation cycles and it was decided to proceed for the scale-up to 80 L for
51
the second submission batch for Product Y for Injection USP 10 g/vial. Though, after unloading 7% of
vial breakage was still verified.
In order to completely overcome the problem of vials breakage, additional tests were performed by
R&D Jordan and a new modified lyophilisation cycles were applied in 20 L trial batch (Trial #3 and Trial
#4).
First submission batch
During the first submission batch it was verified around 90% of the vials were broken. The applied
cycle was following:
Phase No. Process-Phase Phase Time (h: m) Set Temp. (ºC) Vacuum (µbar)
1. Loading (Precooled shelves) --:-- +5
OFF
2. Freezing
06:00 -50
04:00 -50
3. Primary Drying (Sublimation)
06:00 -20
770
38:00 -20
06:00 -10
04:00 -10
04:00 0
23:00 0
03:00 +10
04:00 +10
02:00 +15
02:00 +15
4. Secondary Drying (Desorption)
02:00 +20
520
03:00 +20
03:00 +25
08:00 +25
02:00 +35
52
12:00 +35
Total Cycle Time 132:00 hours
Table 2. First submission lyophilisation cycle parameters.
The main reason for vial breakage was attributed to the fast-freezing rate applied – ramp down to -50
ºC in 6 hours - a factor that is especially important to take in consideration when applied on high
concentrated solutions and large fill volumes, as the case of Product Y 10 g/vial. Following this
investigation, a trial batch with slower freezing rate was applied, Trial #1.
Trial #1
The main goal was to elongate the freezing time from 6 hours to 12 hours. The first 28 hours of the
cycle applied on Trial #1 were considered sufficient to evaluate the performance of the cycle in terms
of containers breakage.
Phase No. Process-Phase Phase Time (h: m) Set Temp. (ºC) Vacuum (µbar)
1. Loading (Precooled shelves) --:-- +5
OFF
2. Freezing
12:00 -50
04:00 -50
3. Primary Drying (Sublimation)
06:00 -20
770
06:00 -20
Total Cycle Time 28 hours
Table 3. First R&D trial lyophilisation cycle parameters.
The Trial #1, was achieved successfully on the R&D-Lyophilizer, the containers did not show any
evidence of breakage. Later on, this cycle was applied at Hikma Farmacêutica Portugal and signs of
cracks on the vials surface started to appear after 28 hours of the cycle. To have a better insight, it
was decided to proceeded up to 69 hours and after the unloading, 167 out of 257 vials were broken,
representing 66% of the total batch. Despite the elongation time could slightly decrease the
percentage of broken vials, a new development trial was made to overcome the problem.
Trial #2
The following modifications were introduced on Trial #2:
53
• Adding a hold step for 1h at -10 ºC, the super-cooling temperature, in order to help arrange
the crystals;
• Adding a hold step for 3h at -20 ºC (Product Y freezing point);
• The terminal freezing point was modified to -45 ºC instead -5 ºC;
• Reducing the holding time at -45 ºC from 4h to 3h;
• Decreasing the ramp-up (from -45 ºC to -20 ºC) time from 6h to 3h to enhance the drying of
the cake.
The 3rd and 4th step were introduced to reduce the time at the freezing stage which will minimize the
pressure exerted on the wall of the vials. All the modifications were due to prevent sudden expansion
due to initial crystallization and reduce the time of the product at frozen state which would minimize
the physical stress exerted by the frozen structure on the vial.
Phase No. Process-Phase Phase Time (h: m) Set Temp. (ºC) Vacuum (µbar)
1. Loading (Precooled shelves) --:-- +5
OFF
2. Freezing
02:00 -10
01:00 -10
01:00 -20
03:00 -20
03:00 -45
03:00 -45
3. Primary Drying (Sublimation)
03:00 -20 770
38:00 -20
520 06:00 -10
04:00 -10
04:00 0
420
23:00 0
03:00 +10
04:00 +10
02:00 +15
02:00 +15
4. Secondary Drying (Desorption) 02:00 +20 310
54
03:00 +20
03:00 +25
08:00 +25
02:00 +35
12:00 +35
Total Cycle Time 131:00 hours
Table 4. Second R&D trial lyophilisation cycle parameters.
After vials unloading, two vials out of the 213 vials were found broken representing less than 1% of the
total batch indicating that the applied cycle modifications were successful in improving vial breakage
during freezing.
Regarding the R&D-Lyophiliser successfully results, an 80 L submission batch was produced with the
following modifications:
• Holding step for 1h at -12 ºC, the super-cooling temperature;
• Holding step for 4h at -20 ºC, the Product Y freezing point;
• Increasing the ramp-down (from -20 ºC to -45 ºC) time from 3h to 3h and half.
When the lyophilisation cycle was finished it was noticed 7% of vials breakage, and compared with the
first submission batch in which 90% of broken vials were verified, the new cycle did not completely
solve the problem of vials breakage.
In addition, the reconstitution time was longer suggesting that the lyophilised cake was not totally dried
forming a lump upon the addition of reconstitution water complicating the dissolution.
Trial #3
The following modifications were purpose on Trial #3:
• Slow ramp down to -18 ºC in 5h;
• Adding a hold step at -18 ºC for 6h;
• Slow ramp down to -45 ºC in 5h because is expected with a slower freezing rate, large
crystals;
• Holding for 4 hours at -45 ºC;
• Addition of annealing step by increasing the temperature to -18 ºC in 2 hours, holding for 4
hours at -18 ºC and then lowering the temperature down to -40 ºC in 3 hours. This would help
rearranging the crystals and obtaining a more homogeneous frozen structure;
55
• Start the primary drying at -10 ºC instead of -20 ºC to help in better and faster primary drying;
• The vacuum applied in primary drying was increased to 100 µbar to enhance the water vapour
sublimation from this large volume cake.
After unloading, 55 % of the vials were found broken showing a non-effective cycle, which did not
solve the minor percentage of 1% of vial breakage (trial #2) and the 7% in the subsequent 80 L
submission batch. More particularly, the addition of the annealing step was not positively.
Trial #4
The modifications involved in the development of this cycle were:
• Removing the annealing step during product freezing stage;
• Addition of a hold step for 6h at -20 ºC, the super-cooling temperature at which the crystals
formation and arrangement start;
• Modification of the terminal freezing point to be -45 ºC instead of -40 ºC to ensure that the
product is reaching the freezing point;
• Reducing the Holding time at -45 ºC from 4h to 3h to reduce the time at the frozen state which
will reduce the pressure exerted on the wall of the vial;
• In the sublimation phase, addition of a holding step for 6h at -30 ºC prior to further increase in
temperature to ensure that the cake is completely sublimed before giving further energy.
56
Phase No. Process-Phase Phase Time (h: m) Set Temp. (ºC) Vacuum (µbar)
1. Loading (Precooled shelves) --:-- +5
OFF
2. Freezing
04:00 -20
06:00 -20
05:00 -45
03:00 -45
3. Primary Drying (Sublimation)
00:30 -35
100
03:00 -35
03:00 -30
06:00 -30
05:00 -10
18:00 -10
04:00 0
27:00 0
03:00 +10
05:00 +10
4. Secondary Drying (Desorption)
00:30 +10
300
03:00 +20
04:00 +20
01:00 +25
12:00 +25
01:00 +35
19:00 +35
Total Cycle Time 133:00 hours
Table 5. Fourth R&D trial lyophilisation cycle parameters.
In September 2012 a batch with the modifications was manufactured. After vials unloading and 100%
visual inspection, no vials were found broken. This indicates that the applied lyophilisation cycle was
57
successful by solve the problem of vials breakage. Thereafter, the reconstitution time was evaluated
and it was out the specifications.
Forthwith, changes in the optimized cycle (having as reference the cycle applied in trial #4) were
successful in enhancing the mentioned product characteristics: crystallization, cake dryness, water
content and reconstitution time. The changes applied are not anticipated to cause vial breakage since
the phenomenon is directly affected by freezing stage and primary drying. Moreover, this cycle was
challenged by loading 100 mL filled vials along with 60 mL ones and no vials breakage was observed
by the end of the cycle.
Based on the results obtained from all the trials, scale-up and submission batches, it was
recommended to proceed with the preparation of the submission batch of Product Y 10 g/vial applying
the following lyophilisation cycle:
Phase No. Process-Phase Phase Time (h: m) Set Temp. (ºC) Vacuum (µbar)
1. Loading (Precooled shelves) --:-- +5
OFF
2. Freezing
06:00 -20
08:00 -20
06:00 -45
03:00 -45
3. Primary Drying (Sublimation)
00:30 -35
100
03:00 -35
03:00 -30
06:00 -30
05:00 -10
18:00 -10
04:00 0
35:00 0
4. Secondary Drying (Desorption)
04:00 +25 100
00:30 +25
50 16:00 +25
01:00 +35
58
19:00 +35
Total Cycle Time 138:00 hours
Table 6. Final Lyophilisation cycle parameters to be applied for FDA submission.
After all data collection, Hikma Farmacêutica wrote, in 2012, a Data Compilation Report for the
submission of Product Y for Injection, USP 5 g/vial and Product Y for Injection, USP 10 g/vial where
the critical process steps in the manufacturing process were evaluated according to the Risk
Assessment (Appendix 2).
The compounding of the bulk solution formula, bioburden reduction filtration through a 1.2 µm filter,
final filtration through a 0.2 µm filter, filling, lyophilisation and inspection were accepted in October,
2018 when the FDA gives the approval for the manufacturing process for Product Y for Injection, USP
5 g/vial and 10 g/vial.
7.4. Practical Case
After the review of all data collected about Product Y and an exhaustive investigation of the life cycle it
was possible to confirm that the problem was in the freezing phase similar what happened in the past
with the 10 g/vial form. In the freezing phase, it happens quickly the icing of the lyophilised cake and
this step causes an expansion in the cake volume.
In order to improve and solve the cause of this type of breakage during lyophilisation attributed to the
freezing phase it was proposed to adjust the recipe (LYC018) of Product Y 5 g/vial to become similar
to the freezing step of the Product Y 10 g/vial, already improved in the past, in which the freezing
pattern is gradual rather than shocking the matrix sudden and with very low temperature.
To adapt the recipe of Product Y 10 g/vial in the recipe of Product Y 5 g/vial it was necessary to
perform a Risk Assessment where all the risks were evaluated.
59
Approved 10 g/vial lyo cycle Approved 5 g/vial lyo cycle
Time
(hr:min)
Temp
(°C)
Vacuum
(µbar)
Time
(hr:min) Temp (°C)
Vacuum
(µbar)
Freezing
6:00 -20
-
8:00 -20
6:00 -45 5:00 -50
-
3:00 -45 4:00 -50
Table 7. Approved freezing phase parameters.
In terms of the lyophilisation cycle, Product Y, USP 10 g/vial recipe presentation is considered the
worst-case, having the double of the fill volume (and the same container than in Product Y 5 g/vial)
and higher stress and impact on the Primary Package, namely the vial itself. However, the 10 g/vial
presentation shows not suffer from breakage as the 5 g/vial.
Upon reviewing the Lyophilisation Cycle for both strengths; it was found out that the 10 g/vial has a
gradual freezing pattern in the cycle, while the 5 g/vial lacked this, but rather it was a sudden and
much fast freezing without any intervals.
It was extrapolated that by applying the same freezing rates and pattern to the 5 g/vial this vial
breakage will be eliminated, and the frozen matrix will behave similarly to the 10 g/vial (as both share
exactly the same formulation).
Since both formulations share the same bulk solution, container closure system – both are filled in a
100mL molded vials- and they only differ in the fill volume it was concluded during the Risk Analysis
that the adaption of the freezing recipe of Product Y 10 g/vial in the recipe of Product 5 g/vial has no
impact in the quality of the product and manufacturing process.
The modifications performed in the freezing step intend to decrease the impact of lyophilisation on the
rejects quantity. No changes were made to the rest of the recipe, primary and secondary drying, for
Product Y for Injection, USP 5 g/vial.
60
Approved 10 g/vial lyo cycle Approved 5 g/vial lyo cycle Improved 5 g/vial lyo cycle
Time
(hr:min)
Temp
(°C)
Vacuum
(µbar)
Time
(hr:min)
Temp
(°C)
Vacuum
(µbar)
Time
(hr:min)
Temp
(°C)
Vacuum
(µbar)
Loading - +5 - - +5 - - +5 -
Freezing
6:00 -20
-
6:00 -20
-
8:00 -20 8:00 -20
6:00 -45 5:00 -50
-
6:00 -45
3:00 -45 4:00 -50 3:00 -45
Primary
Drying
00:30 -35
100
2:00 -20
770
2:00 -20
770
3:00 -35 12:00 -20 12:00 -20
3:00 -30 4:00 -10 4:00 -10
6:00 -30 4:00 -10 4:00 -10
5:00 -10 3:00 0 3:00 0
18:00 -10 19:00 0 19:00 0
4:00 0 3:00 +10 3:00 +10
35:00 0 2:00 +10 2:00 +10
2:00 +15 2:00 +15
2:00 +15 2:00 +15
Secondary
Drying
4:00 +25 100 2:00 +20
520
2:00 +20
520
00:30 +25
50
3:00 +20 3:00 +20
16:00 +25 2:00 +25 2:00 +25
1:00 +35 3:00 +25 3:00 +25
19:00 +35 2:00 +35 2:00 +35
8:00 +35 8:00 +35
Table 8. Lyophilisation cycle parameters comparison for Product Y 5 g/vial (Current and Improved
Freezing Phase) and 10 g/vial.
Every time an ANDA holder needs to do a modification in a manufacturing process already approved it
is necessary to report to the authorities. Depending on the type of change, the applicant must notify
FDA about the change in a supplement or by inclusion of the information in the annual report to the
application.
There are two types of moderate change:
61
- One type of moderate change requires the submission of a supplement to FDA at least 30 days
before the distribution of the drug product made using the change “Changes Being Effected in 30
Days” ("CBE-30").
- CBE-0 is a moderate change for which distribution of the product can occur when FDA receives the
supplement.
To do the modification in the lyophilization recipe it was necessary to submit a "Changes Being
Effected" ("CBE-0") for FDA.
After all the changes were made, an evaluation of 100% inspection was done and compared with
another PV batch of Product Y 5 g/vial that was lyophilised using the approved freeze-drying cycle
before the improvement.
APPROVED LYO CYCLE AS
PER LYC018
IMPROVED FREEZING
PHASE CYCLE
Total vials 4779 4980
Total of broken/cracked vials rejected 652 36
% of total of broken/cracked vials rejected 13.6 0.7
Table 9. Side-by-side Inspection results of batch of Product Y for Injection USP 5 g/vial.
From the evaluation of the analytical data presented from the manufactured batches of Product Y for
Injection USP 5 g/vial, no negative impact is observed on the product quality after the freeze-drying
phase cycle improvement. As it can be observed from Table 9, it was noticed a decrease of cracked
vials from 13.6% to 0.7% in the capping loading process.
7.5. Line Transfer and Scale-up for Product 5 g/vial
– a Process Validation
After the successful improvements, it was further suggested a Line Transfer from Line 1 to Line 9 and
a scale-up from 170 L to 310L batch size of Product Y for Injection USP 5 g/vial for US Market. For
that, it was necessary to perform a Process Validation Protocol to define the activities and sampling
requirements intended to evaluate and validate the scale-up and the Line Transfer.
In order to validate the scale-up process, three Process Validation (PV) batches of 310 L of Product Y
for Injection USP 5 g/vial were manufactured at Line 9 in Hikma Farmacêutica S.A. One PV batch was
62
for submission purpose and, upon approval of the process two additional PV batches were
manufactured, in order to complete the process validation activities.
After the filling the PV batches, the vials were submitted to 100% inspection for particles and defects.
All activities required for full qualification of facilities, equipment and systems to be used on the
process, were previously made and approved. All batches were tested using validated analytical
methods and, raw-materials and primary package components were analysed and released before
production.
Hikma Farmacêutica, S.A validation program includes the validation of processes used in
manufacturing pharmaceutical products. In the case of Product Y for Injection USP 5 g/vial, this
includes compounding of the bulk solution, pre-filtration through a 1.2µm filter, filtration through
sterilizing 0.2µm filter, filling, lyophilisation, stoppering, capping and 100% visual inspection. The
filtered bulk solution was filled into a 100 mL vial presentation.
The critical process steps in the manufacture of Product Y for Injection USP 5 g/vial were evaluated
based on Risk Management Tools (FMECA) as per the following rational:
7.5.1. Process Evaluation Activities and Results
The following activities were performed during the production of all validation batches in order to
define certain process details and to guarantee that the manufacturing process is suitable for the
production of Product Y 5 g/vial at Line 9 with a batch size of 310 L.
Evaluation of quality of the compounded bulk solution
The range of mixing speeds and processing durations used were evaluated during compounding to
assure that complete dissolution occurs under all approved compounding conditions, dissolution
should be complete, and the solution should be uniform. Different speeds were used in the three
batches based on previous experience with similar products compounded using the same tanks.
After the final mixing step was complete, samples from the top and bottom of the compounding vessel
were taken for physiochemical analysis and from the bottom for microbiological analysis.
It was observed a high dissolution time during the compounding, studies are being performed by R&D
department in order to optimize the compounding step and the compounding parameters needs to be
re-evaluated on the next commercial batches.
Evaluation of effects of elapsed compounding (Bulk holding time)
63
Although this holding time was previously established as 24h in the preparation tank, this holding time
was re-evaluated at cold conditions (8ºC – 12ºC) until 48h after transference process.
The bulk solution of one batch presentation was evaluated for the maximum bulk holding time in the
preparation tank. Samples were taken from the bottom of the preparation tank at 0h, 18h, 24h and 48h
after the completion of transference activities, for IPC and HPLC and samples for microbiological
testing were collected at 0h, 24h and 48h.
Based on results obtained from one batch, it can be concluded that a bulk holding time of 48 hours
(with transference activities) has no negative impact on the product quality and the product from both
presentations can stay up to 48h in the transference tank from the end of compounding until the end of
filling.
Evaluation of the effects of initial set up (Dead Volume) and line stoppages
An initial quantify of filled units is typically discarded as part of initial line set up. The purpose of this
study was to establish the minimum amount of product that needs to be discarded prior to start filling.
For this reason, samples were collected from each batch before staring the filling, and the system was
purged.
The results obtained on second batch, Product Y (%), pH and related substances are within
specifications and aligned with the finished product results. Therefore, it can be concluded that no
impact is observed on product quality after purging the system with 300 mL and no further actions are
required.
To establish acceptable line stoppage duration and determine the minimum quantity of units to be
discarded prior to re-start filling, one line stoppage of a minimum of two hours was incorporated into
the filling. Samples were collected for analysis and it was observed that the first vials from stage#1
were already within specifications and aligned with finished product results. Therefore, it can be
concluded that line stoppages up to 2h have no impact on the quality of the product, and no further
actions are required.
Evaluation of the effects of filtration and filling on the quality of the
compounded solution
To evaluate the filtration and the filling uniformity, filled units were collected from the beginning, middle
and end of the filling process for testing and every hour for visible particles. The beginning of the
process is defined as a point in which the filtered bulk solution is no longer discarded and line flush
and set up activities have been completed. Sampling activities are designed to validate the entire
filling process.
64
From the manufacturing process and in-process controls it can be concluded that all samples met in-
process physiochemical criteria. The filling machine is capable of filling 30.5 mL in 100 mL vials within
the specified parameters for the 310L batch size and the filling process does not affect the product
quality. Additionally, results of beginning, middle and end obtained after lyophilisation are within the
acceptance criteria hence assuring the quality of the filled solution.
Evaluation of the filling machine speed
The maximum and minimum filling machine speed were challenged in order to set a filling speed
range. Filled units were collected at the minimum and maximum filling process speed from the filler
and from the evaluation performed, it can be concluded that the filling machine is capable of filling
30.5mL in 100 mL vials sizes within specification, with a filling machine speed between 40 vials/min
and 70 vials/min.
Evaluation of the holding time between end of API addition and beginning of
Lyophilisation cycle
This holding time is covered by bulk holding time studies in the transference tank performed and, the
holding time between the end of API addition and the beginning of lyophilisation cycle, (freezing
phase) for Product Y for Injection USP, 5 g/vial was 26h 35min for one batch. However, the holding
time between the end of API addition and the beginning of lyophilisation cycle (freezing phase)
already evaluated and considered validated was 24h, consequently the holding time was exceeded
in 02h 35min. Based on the investigation performed, it can be concluded that the present deviation
has no negative impact on the quality of the product of this batch, since from finished product
analytical and microbiologic results are within specification and no out of trend was detected.
Meanwhile, bulk holding time studies performed after transference activities was re-evaluated on
another different batch. This holding time covers the maximum time that the product withstands in
the transference tank (from end of compounding until the end of filling) without affecting the product
quality. The maximum time qualified as part of bulk holding time study is also applicable to determine
the maximum time between end of API addition and the beginning of the lyophilisation cycle as a
worst-case, covering and controlling the total time which the product can stay in the liquid state.
Evaluation of the quality of the Finished Product
Capped vials from the lyophilised product were also taken from the beginning, middle and end of the
process validation batches for physiochemical and microbiological testing as per finished product
criteria. With the results analysis of all batches, it can be concluded that the manufacturing process for
65
the 310L batch size of Product Y for Injection USP, 5 g/vial is capable to produce consistently a
product complying with defined specifications.
Evaluation of the effects of Lyophilisation on the quality of the filled vials and
lyophilisation uniformity
The process validation batches were lyophilised as per lyophilisation cycle in SOP#LYC018.
Lyophilizer from Line 9 was considered the worst case from a loading perspective because the loading
and unloading at Line 9 is automatic while in Line 1 is manual despite reduces aseptic handling. Both
lines have similar lyophilizers however the controls and also the condensers capacity are different.
To evaluate the quality and uniformity of lyophilisation, vials were collected from the freeze-dryer for
analysis (appearance of the cake, reconstitution time and water content) from the indicated positions
presented on Figure 15 and properly labelled accordingly to correlate the results with the position in
the lyophilizer.
Notes:
a) Each batch will fill approximately
10 shelves per lyophilizer.
b) Samples from three shelves per
lyophilizer should be
collected/tested.
c) If the last shelf is not fully loaded
collect samples from the previous
one.
d) All vials must be capped
(manually or using the capper).
Figure 18- Loading positions for PV batches and samples collection.
Based on the results evaluated from samples taken from several lyophilizer positions, it can be
observed that all results are within specifications and aligned between them, which means that there is
uniformity within lyophilizer. Additionally, the water content results show that the lyophilisation cycle is
efficient, properly drying the product. Moreover, the water content results for finished product obtained
are well below the acceptance criteria and aligned when compared with the results of the previous
manufactured batches of Product Y for Injection USP, 5 g/vial with 170L batch size in Line 1.
Middle shelf
First shelf
Last Shelf
66
Despite the lyophilizer from Line 1 and Line 9 are different as also the controls and the capacity it was
possible to confirm with the results from this evaluation that there is uniformity within lyophilizers.
Evaluation of 100% inspection
Lyophilised vials from the process validation batches were submitted to 100% visual inspection for
defects and particles after being capped. The 100% inspection is executed using the automatic
inspection machine. From the results, it can be concluded that the percentage of total number of vials
rejected with defects is between 0.1% and 0.6% of the total number of vials submitted to capping and
100% inspection showing all results are conforming.
Successful process validation studies provide evidence that a defined manufacturing procedure will
consistently yield product meeting its pre-determined specifications and quality characteristics. Each
activity is described in detail as are sampling requirements needed to complete the evaluation.
From the manufacturing process and in-process controls (beginning, middle and end) and from
finished product testing and side by side comparison (Attachment 4), it can be concluded that the
process is capable of producing a product within specifications with scale up from 170L to 310L and
line transfer from Line 1 to Line 9.
After the lyophilisation during the PV activities for the Line Transfer and scale-up it was analyzed the
number of broken vials (Table 11).
B# 1 B# 2 B# 3 B# 4
% of total of broken/cracked
vials rejected 0,04% 0.04% 0,08% 0,2%
Table 11. Percentage of cracked vials reject after Scale-up and Line Transfer.
After scale-up and Line Transfer activities it was possible to verify that all changes done before in the
lyophilisation cycle were successful and without negative impact in the number of broken vials.
Additionally, the quality of the final product was kept. In short, the results shown that we are facing a
robust and validated process who is reproducible in Line 9 with a batch size of 310 L and possible to
launch in the market with high quality.
67
Figure 19. Life cycle of Product Y for Injection after Launching.
7.6. Launching
Fill Volume Price
Product Y 5 g/vial 10.52 €/vial
Product Y 10 g/vial 20.45 €/vial
Table 12. Launching for Product Y for Injection USP.
The Product Y 10 g/vial is valued as 20.45 € per vial and the presentation of Product Y 5 g/vial as
10.52 €/vial.
After the improvement made in Product Y 5 g/vial it was possible to reduce the percentage of
breakage vials from 13.6% to 0.7% which means a decrease in 12,9% of breakage vials per batch.
The theoretical value per batch is as described in Table 12.
Line Minimal Quantity (units) Maximum Quantity (units)
1 5 573 5 573
9 10 163 10 163
Table 13. Approved Batch Sizes for Product Y for Injection, USP 5 g/vial.
After all the modifications, it was measured the percentage of good vials per theoretical batch that was
possible to rise up and calculated the value possible to monetize per batch: 7 563€ for Line 1 and
Launching(Product Y)
5 g/vial
10 g/vial
No problemsdetected
Validatedprocess
2019
High BreakageRate
US Market
Problem: FastFreezing
New LyophilizationRecipe adapted
CBE 0
Line Transferand Scale-up
Line
1
Line
9CBE 30
68
13 792€ for Line 9. In one year if we produce 25 batches in Line 1 and 25 batches in Line 9 it is
possible to increase for 533 875€ per year.
For the future, one Line will be dedicated for Product Y and if in half-year (26 weeks) it is produced
Product Y 5 g/vial two times per week in Line 9, for example, with this modification it will be possible to
improve in 717 184€.
The values represented above do not represent a significant impact for the company. However, with
the improvement of Product Y 5 g/vial it was possible to validate the manufacturing process and
consequently launch the product for the US Market.
8. Conclusion
Before a drug product reach the market and start to be routinely manufactured, it passes through
several steps, from R&D to normal production. During pharmaceutical development, the product and
the manufacturing process are designed to manufacture a product, which is intended to be use in
accordance with regulatory authorities’ requirements and specifications.
When a product and his process are developed, several activities need to be evaluated to ensure that
the process can be reproducibly and consistently to obtain a product with good attributes and desired
quality.
Technology transfer implicates transfer of product and process knowledge to achieve the
manufacturing process for commercialization and it includes all the activities required since the
pharmaceutical development (R&D) to production, for new products, or from one manufacturing site to
another.
Process validation is part of technology transfer and is applied to demonstrate that the manufacturing
process developed, operated within established parameters, can consistently deliver the intended
product. For a successful process validation and consequently technology transfer and scale-up, a
proper correlation between critical material attributes, critical quality attributes of the products and
critical process parameters of the equipment must be well-established. While there are numerous
inputs, outputs and controls correlated it is important to have a systematic tool who understands the
connection between product and process, based on quality risk management, to identify and to
evaluate the process validation activities to be performed during technology transfer.
This includes establishing strong quality management systems, obtaining appropriate quality raw
materials, establishing robust operating procedures, detecting and investigating product quality
deviations and maintaining reliable testing laboratories. It helps to prevent the occurrence of
contaminations, mix-ups, deviations and failures and assures that the drug products manufactured
meet their quality standards.
69
Some manufacturing procedures, as the lyophilisation of products, have a panoply of controls and
parameters since R&D until the normal routine production, making it more difficult to perform.
When a lyophilised product is manufactured, an appropriate lyophilisation cycle should be developed.
During R&D the cycle development is done however adjustments in cycle times and parameters may
occur when transferring from R&D freeze dryer to an industrial lyophiliser.
In 2019, Hikma Farmacêutica started to produce a new product already submitted to the authorities
and approved by them: Product Y for Injection USP 5 g/vial and Product Y for Injection USP 10 g/vial.
During the PV activities it was noticed a high percentage of broken vials of Product Y 5 g/vial
becoming a process non-robust and invalidated. For the improvement of manufacturing process of this
product it was necessary to do an investigation through the life cycle of this product.
The root cause of the breakage problem was attributed to the fast-freezing rate applied during freezing
phase on the lyophilisation cycle. The fast-freezing makes the product to expand suddenly without
allowing the crystals, for enough time, to be shaped according to the vial size creating pressure on the
edges and causing the bottomless breakage of the glass.
During the investigation it was observed that the problem with the breakage rate was noticed in the
past with the 10 g/vial form and it was purposed some modifications on the freezing phase.
In order to optimize the lyophilisation cycle of 5 g/vial form and based in a Risk Assessment it was
purposed to adapt the freezing recipe of 10 g/vial in the recipe of 5 g/vial. Changes were made and
the improvement of lyophilisation cycle, namely, the freezing phase was successfully and it was
possible to decrease the percentage of vial breakage from 13.6% to 0.7% of vials per batch becoming
a validated process and therefore open the possibility to launch the product in the US Market.
After the successful improvement, a Line Transfer from Line 1 and Line 9 and a scale-up to 310 L
were proposed and it was necessary to perform a Process Validation where the critical steps were
analysed. The PV results showed no negative impact resulting in an effective Line Transfer and scale-
up.
The development and optimization of a manufacturing process of a Lyophilised product can be
sometimes a “tricky and complex” question. The development phase, of routine production as wells as
the scale-up and technology transfer of lyophilisation processes, remain challenging.
In short, it is also important to have an effective tool to collect data to be able to improve based on
past experiences.
70
8.1. Future work
The manufacturing of injectable pharmaceutical product, in particular lyophilized products, is a
complex process. There are many different operations with several parameters and each unit
operation must be properly validated and controlled in order to assure obtaining a sterile drug product
in accordance with the specifications and regulatory authorities.
Nowadays, many pharmaceutical companies dismiss many tests with trial and error process and
consequently time and money expenditure. For the future, mainly at Hikma, it will be interesting if
other applications should be considered as an opportunity to ease and improve the process.
Process Analytical Technology (PAT) is a mechanism to provide a higher degree of process control I
which analyzes, designs and control pharmaceutical manufacturing processes through the
measurement of Critical Process Parameters (CPP) which affect Critical Quality Attributes (CQA). It
allows real-time monitoring and control to adjust the processing conditions so that the output remains
constant, being more efficient in testing while at the same time reducing over-processing, enhancing
consistency and minimizing rejects. PAT can be used in the pharmaceutical industry, both for process
validation activities and routine process control.
Reduction of time in production cycles, avoid waste during the manufacturing process, enables real-
time process control facilitating continuous processing, improves efficiency and variability
management are some of the advantages of PAT which can represent a benefit over other
pharmaceutical companies that do not apply this technology.
Although, the implementation of a PAT system is a large financial investment, which requires skilled
and skilled human resources, process changes, equipment modification and the operation of the
pharmaceutical units.
Hikma Farmaceutica, S.A. does not make use of PAT tools in manufacturing processes. The
processes are carried out using the most traditional methods and the production is done in batches.
However, it would be interesting in future terms, to implement PAT and develop a more in-depth study
of the following aspects:
- Trend analysis of the results and deviations observed during commercial batches and
validation batches to identify the most frequent problems and identifying improvement
opportunities and their potential benefits;
- Determining causes that lead to the current situation and possible opportunity areas;
- Defining the variables that may impact on the process and the extent they do;
- Running experimental trials to control or optimize variables;
- Defining measures that allow us to maintain consistently trough time achieved improvements;
- Examination about which could be the most necessary PAT tools to implement at Hikma
Farmacêutica.
71
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74
10. Appendix
Attachment 1- Risk Assessment and Process Validation Approach of the batch for submission of Product Y for Injection USP 5 g/vial and Product Y for
Injection USP 10 g/vial (First Submission Batch).
Process Risk Assessment
Evaluation (ID) Validation Approach Rationale Necessary validation activities
Evaluation of quality of the
compounded bulk solution From A to E
The two presentations have the same bulk formulation but different batch sizes
(40 L and 80 L) One batch of each size
Evaluation of the effects of elapsed
compounding (Bulk hold time) N/A
The two formulations have the same bulk formulation. The batch size parameter
has no impact for this evaluation One batch of any presentation
Evaluation of the effects of filling on
the quality of the compounded
solution
E The two presentations will be filled in 100 mL vials with different fill volumes One batch of any presentation
Dead Volume and Line Stoppages
(up to 2h) N/A
The two presentations will be filled in 100 mL vials. The fill volume has no impact
for this evaluation
One batch of Product Y for
Injection USP 10 g/vial
Holding time between end of API
addition and beginning of
lyophilisation cycle (loading phase)
N/A
The two presentations will have different holding time between end of API
addition and beginning of Lyophilisation cycle due to the different batch size.
Since the vials will be loaded in cool shelves (5 ºC) it could be considered for the
Holding time study the actual time the vials are loaded
One batch of any presentation
75
Evaluation of the effects of
lyophilisation on the quality of the
filled Vials
G
The two presentations will be filled in 100 mL vials with different fill volumes and
different lyophilisation cycles will be used. After lyophilisation the lyophilised FP
specifications are not the same
One batch of any presentation
MICRO Validation N/A
The two presentations have the same bulk formulation. Samples for Sterility and
Endotoxins Validation should be collected for (FP) and samples for Bioburden
and Bacterial Endotoxins should be collected for Bulk solution
One batch of any presentation
Filter Validation N/A The two presentations have the same product formulation One batch of any presentation
76
Attachment 2 - Risk Assessment and Data Compilation Approach with the batches used to qualify the Product Y for Injection, USP 5 g/vial and 10 g/vial
Process
Risk
Assessment
Evaluation (ID)
Validation Approach Rationale
Necessary
validation
activities
Batches evaluated
Evaluation of quality of
the compounded bulk
solution
From A to E The two presentations have the same bulk formulation but
different batch sizes (40 L and 80 L)
One batch of each
size
One batch of Product Y for Injection USP 5
g/vial
One batch of Product Y for Injection USP 10
g/vial
Evaluation of the
effects of elapsed
compounding (Bulk
hold time)
N/A The two formulations have the same bulk formulation. The
batch size parameter has no impact for this evaluation
One batch of any
presentation
One batch of Product Y for Injection USP 10
g/vial
Evaluation of the
effects of filling on the
quality of the
compounded solution
E The two presentations will be filled in 100 mL vials with
different fill volumes
One batch of any
presentation
One batch of Product Y for Injection USP 5
g/vial
One batch of Product Y for Injection USP 10
g/vial
Dead Volume and Line
Stoppages (up to 2h) N/A
The two presentations will be filled in 100 mL vials. The fill
volume has no impact for this evaluation
One batch of
Product Y for v USP
10g/vial
One batch of Product Y for Injection USP 10
g/vial
Holding time between
end of API addition and
beginning of
lyophilisation cycle
N/A
The two presentations will have different holding time
between end of API addition and beginning of
lyophilisation cycle due to the different batch size. Since
the vials will be loaded in cool shelves (5 ºC) it could be
considered for the Holding time study the actual time the
One batch of any
presentation
One batch of Product Y for Injection USP 5
g/vial
One batch of Product Y for Injection USP 10
g/vial
77
(loading phase) vials are loaded
Evaluation of the
effects of lyophilisation
on the quality of the
filled Vials
G
The two presentations will be filled in 100 mL vials with
different fill volumes and different lyophilisation cycles will
be used. After lyophilisation the lyophilised FP
specifications are not the same
One batch of any
presentation
One batch of Product Y for Injection USP 5
g/vial
One batch of Product Y for Injection USP 10
g/vial
MICRO Validation N/A
The two presentations have the same bulk formulation.
Samples for Sterility and Endotoxins Validation should be
collected for (FP) and samples for Bioburden and
Bacterial Endotoxins should be collected for Bulk solution
One batch of any
presentation
One batch of Product Y for Injection USP 10
g/vial
Filter Validation N/A The two presentations have the same product formulation One batch of any
presentation
One batch of Product Y for Injection USP 5
g/vial
78
Attachment 3
Product Y for Injection, USP 5g/vial
Flow Diagram for Batch Preparation- Line 9
Excipient 1
Mix until complete homogenization. Keep the temperature (8 – 12ºC).
Check the pH of the solution (2.5 – 4.5).
Mix slowly and under vigorous agitation
assuring its complete incorporation.
Sparging with filtered nitrogen until dissolved
oxygen < 1 ppm.
Decrease the agitation and agitate until
complete dissolution and Sparging between
Stainless Steel
Sparging with filtered nitrogen until dissolved
oxygen ≤ 0.25 ppm.
Product Y, USP
Product Y, USPP
Adjust volume to 100%.
Mix until complete homogenization.
Sparging with filtered nitrogen until dissolved
Deaereted WFI (8-12ºC)
Deaereted WFI (8 –
Start
Notes: Protect the samples from air
exposure, from light (by wrapping with
aluminium foil) and from heat sources.
End
IPC:
-Description
-pH
-Density
QC:
Deaerate WFI for a
minimum of 20 min
(≤ 20ºC)
Cold WFI
Cold WFI (8 – 12ºC)
20L Stainless Steel
Container
79
Grade C
20mm red
aluminium Flip-off
Secondary packaging
Capping (under UFH – Grade A air
supply)
Grade C
Compounding in SS
Tank
Pallets preparation
100% Visual inspection for defects
Freeze Dryers
Grade A
Frames loader
Grade A
Product filtration by
Grade C
VHP
Chamber
Filling and Stoppering
Grade C
Pre-filtration by 1.2
µm filter
Shipping
Final filter integrity
test before and
Grade C Area
Grade A Room
Unclassified - Warehouse
Unclassified - Packaging
Area
Unclassified - Warehouse
Active and Inactive
Raw Materials
100ml vials, neck
20 mm, Type I,
Moulded, Clear 20mm Lyo Stoppers
Filling size
parts
Frames
Grade D
Attachment 4 – Scale up and Line Transfer evaluation – side by side
A side by side comparison of finished product results was performed between 170L batches manufactured in Line 1 and 310L batch manufactured in Line 9 of Product Y for Injection USP, 5 g/vial in order to evaluate the reproducibility of
results or if any manufacturing constraint related to the scale up process and the line transfer process is observed.
Table A2.1 – Finished Product Results of batches of Product Y for Injection USP 5 g/vial manufactured in Line 1 with 170L batch size vs Line 9 with 310L batch size
Conclusion: From the evaluation of the analytical data presented from the manufactured batches of Product Y for Injection USP 5 g/vial no negative impact is observed on the product quality after scaling up from 170L to 310L batch
size, transferring the manufacturing process from Line 1 to Line 9.
Batch # 1909010.1 Batch # 1909098.1 Batch # 1909099.1 Batch # 1909099.2 Batch # 1901016.1 Batch # 1901015.1 Batch # 1901014.1 Batch # 1901013.1 Batch # 1901012.1
Mfg. Date: January 2019 Mfg. Date: July 2019 Mfg. Date: July 2019 Mfg. Date: July 2019 Mfg. Date: January 2019 Mfg. Date: January 2019 Mfg. Date: January 2019 Mfg. Date: January 2019 Mfg. Date: January 2019
Reference SOP: FPL031 - Rev. G Exp. Date: January 2021 Exp. Date: July 2021 Exp. Date: July 2021 Exp. Date: July 2021 Exp. Date: January 2021 Exp. Date: January 2021 Exp. Date: January 2021 Exp. Date: January 2021 Exp. Date: January 2021
Tests Standard Specifications Results Results Results Results Results Results Results Results Results
1. Description Off-white to tan colored lyophilized powder Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms
2. Identification
2.1. By IR Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms
2.2. By HPLC Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms
3. Water (%) NMT 5.0 2.2 2.9 2.6 2.7 1.8 1.6 1.6 1.7 1.5
4. pH 2.5 to 4.5 3.7 3.2 3.5 3.5 3.3 3.6 3.7 3.7 3.7
5. Uniformity of Content (WV)The Acceptance Value of the first 10
dosage units ≤ L1%0.4% 0.7% 0.9% 1.1% 2.9% 0.7% 0.7% 1.0% 1.7%
6. Bacterial Endotoxins ( USP EU/mg) NMT 0.33 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02 < 0.02
7. Sterility Sterile Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms
8. Particulate Matter
8.1. Sub-Visible particles
Ø ≥ 10 µm NMT 6000 particles/Vial 4053 3460 3673 4280 1640 4660 5300 2900 2860
Ø ≥ 25 µm NMT 600 particles/Vial 73 140 60 93 13 73 93 73 40
8.2. Visible Particles Essentially free from visible particle matter Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms Conforms
9. Color and clarity of constituted solution
9.1. Color (Abs) ≤ 0.125 0.020 0.016 0.012 0.011 0.029 0.018 0.017 0.012 0.012
9.2. Clarity (T %) ≥ 90 100 100 100 100 100 100 100 100 100
10. Assay (%) 90.0 – 115.0 99.3 101.7 102 101.9 97.2 99.6 98.6 101.6 102.7
11. Vancomycin B (%) NLT 90.0 94.5 94.2 94.7 94.7 94.6 94.8 94.9 95.0 95.2
12. Related substances (%)
EP Impurity A NMT 4.0 0.47 0.59 0.60 0.60 0.47 0.48 0.56 0.62 0.46
EP Impurity B (B1 + B2) NMT 4.0 2.17 2.7 2.13 2.15 2.41 2.10 2.05 2.01 2.09
EP Impurity C NMT 4.0 0.12 0.14 0.12 0.13 0.13 0.10 0.11 0.11 0.10
EP Impurity D NMT 4.0 0.31 0.33 0.31 0.31 0.35 0.26 0.26 0.28 0.28
Other Individual Unspecified Impurity NMT 3.0 0.70 0.66 0.72 0.76 0.53 0.69 0.65 0.62 0.51
Total Impurities NMT 10.0 5.3 5.6 5.2 5.2 5.3 5.1 4.9 4.9 4.5
13. Heavy Metals (ppm) NMT 30 < 30 < 30 < 30 < 30 < 30 < 30 < 30 < 30 < 30
14. Ethanol Content (ppm) NMT 10000 376 1068 1237 937 1334 1170 1125 1477 1301
15. Reconstitution Time (min) NMT 3 2 1 1 1 1 2 2 2 2
16. Residual Solvents Complies Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1 Complies w ith USP <467> Option 1
17. Elemental Impurities CompliesComplies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH Q3D
Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
Complies w ith USP <232> and ICH
Q3D Option 2b
18. Injections and Implanted Drug products Complies Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1> Complies w ith USP <1>
Product Name: Vancomycin Hydrochloride for Injection, USP
Label Claim: 5.0 g Vancomycin/ vial
170L batch size - Line 1310L batch size - Line 9
