View
3.870
Download
1
Category
Tags:
Preview:
DESCRIPTION
Buy Silica Gel Powder for Silica Gel, Aluminium Oxide, Paper and Flash Column Chromatography us in Pharmaceutical Industries – Bulk Drugs & API, Nutraceuticals, Herbal Extracts products manufacturers, Research Laboratories, Laboratories Chemical Repackaging, Contract Research Laboratories. Column Chromatography is one of the most useful methods for purification & separation (Isolation) of individual desire compound from mixture of unwanted compounds.
Citation preview
Column Chromatography
PRODUCTS:SILICA GEL
ALUMINIUM OXIDE
What is Chromatography?Chromatography has been developed into a new method of
separation of mixture of compounds mainly when they are available in small quantities.
This method is very useful when the components of a mixture have almost the same physical and chemical properties and hence can’t be separated by other usual methods of separations.
The term chromatography means writing in colour (Chroma = Colour & Graphy = To write).
Types of Chromatography
• Paper Chromatography
• Gas Chromatography
• Thin Layer Chromatography
• Solid - Liquid Chromatography (Column Chromatography)1. Gravity Chromatography
2. Flash Chromatography
3. High performance Liquid Chromatography
What is column chromatography?• Column chromatography is one of the most useful methods for
purification & separation (Isolation) of individual desire compound from mixture of unwanted compounds.
• It is often used for preparative applications on scales from micrograms up to kilograms
• It is a solid - liquid technique in which the stationary phase is a solid & mobile phase is a liquid.
• The stationary phase or adsorbent in column chromatography is a solid. The most common stationary phase for column chromatography is Silica Gel, followed by Alumina Oxide.
• The mobile phase or eluent is a liquid. It is either a pure solvent or a mixture of different solvents.
• It can be used for molecules whose molecular weight is < 2000 g/mol
High Performance Liquid Chromatography
Thin Layer ChromatographyThin layer chromatography, or TLC, is a method for analyzing mixtures by separating the compounds in the mixture. (All types of Chromatography works on same principle.)
Uniform layer of Silica Gel coated onto a piece of glass, aluminium or rigid plastic plates are being used for analysis.
The mobile phase moves through the stationary phase and carries the components of the mixture with it.
Different components travel at different rates.
Application:
To determine the number of components in a mixture,
Identification of compounds, and
Purity of a compound.
Thin Layer ChromatographyIt is used in Analytical study to analyzed microgram (0.000001 g) quantity samples and it takes around 5-10 minutes for result.
Size available:
1. Silica Gel G for TLC (With Binder)
2. Silica Gel H for TLC (Without Binder)
3. Silica Gel GF 254 – with fluorescent indicator
4. Silica Gel HF 254 with fluorescent indicator.
In Thin Layer Chromatography fluorescent is added in stationary phase to analyze substances which are colourless. This we can not see with our naked eyes.
When plate will expose to UV light then it will Glow except the spots of substances & spot will look like a dark patch.
Selection of Stationary Phase & Mobile Phase
• Removal of impurities • No. of components to be
separated • Length of the column
used • Affinity differences
between components • Quality of adsorbent
used
Stationary Phase (Silica Gel or Alumina)
• It requires balancing act between solvent & compound’s polarity.
• The compound must also be soluble in water so they are not permanently adsorbed.
Mobile Phase (Solvents)
• Columns are available in glass tubes, stainless steel & vary in length & diameter.
• Resolution of column depends on both diameter & length of adsorbents packed in column.
• Resolution improves with increase in length & reduces with increase in diameter.
• For e.g. 25 gms of adsorbents will provide a better separation in a 1 cm diameter column than 2 cm diameter column.
• Column dimensions - length & diameter ratio (10:1, 30:1 or 100:1)Column
How Scale Up take place
• Analytical Scale: Column Inner Dia - 4,5,6 mm & it is done at milligram scale.
• Semi Preparative: Column Inner Dia - 10, 20, 30 mm & it is done at milligram scale.
• Preparative: Column Inner Dia – 40, 60, 80, 80 mm & it can be done at gram Scale
• Pilot scale: Kg scale (1, 2, 4, 10, 12 Kg)
• Commercial scale: Bulk Quantity (50, 100, 200, 400 Kgs or more)
Methods of Column PackingDry Method :
Add dry silica / Alumina to the column and apply to the bottom of the column. This will compress the silica gel and keep it compressed for the next steps. Packing can be improved by tapping the column.
While applying vacuum; pour solvent in it.
Allow the solvent to move though the column until reaches to the bottom. At this stage vacuum is not require.
Allow 5–6 columns value of solvent to flow through the column to make sure it is complete packed.
Drain the solvent till the solvent level is just even with the surface of the stationary phase
Methods of Column PackingWet Method:
Fill the column about one third with solvent
In a beaker, measure out the required amount of silica / alumina.
In another beaker, take solvent approximately one and a half times the amount of silica / alumina.
Add silica/alumina to the solvent while swirling in small quantity at a time. Use a glass rod to mix the slurry.
Pour some of the slurry into column & allow solvent to drain to avoid overflowing.
Tap the column carefully to encourage bubbles to rise and the silica to settle
Continue to move the slurry to the column until all the silica or alumina is added.
Wash the inside of the column by pouring solvent down the inside edge.
Drain the solvent till the solvent level is just even with the surface of the stationary phase
How does separation take place?
Types of Columns
Gravity Column Chromatography: Solvent is allowed to move down the column by gravitational forces.
Flash Chromatography:Solvent is pushed down the column by positive air pressure
ApplicationSeparation of mixture of compounds
Purification process
Purification of Phytochemical
Isolation of metabolites i.e. Small molecules
Estimation of drugs
Process DevelopmentPurify Natural compounds
To separate active component from Plant material
Herbal Extraction
Adsorbents used in chromatography method
• Silica gel (SiO2) and alumina (Al2O3) are two adsorbents commonly used for column chromatography.
• These adsorbents are sold in different mesh sizes such as 60-230 mesh, 100-200 mesh, 200-400 mesh & tailor made.
• Adsorbent particle size affects how the solvent runs through the column.
• Smaller particles (higher mesh size i.e. 230-400 mesh) are used for flash chromatography & larger particles (lower mesh size i.e. 60-120/60-200) are used for gravity chromatography.
Difference between Normal Phase & Reverse Phase Chromatography
Normal Phase Chromatography
• It uses a polar stationary phase and a non-polar (low Polarity Solvents) mobile phase.
• Non-polar compounds elute faster than polar compounds.
• When we increase polarity of mobile phase elution time will increase.
• It can not be reused / reproducible
• Mobile phase are non polor i.e. IPA, hexane, dichloromethane, chloroform, ethyl ether, and isopropyl alcohol (IPA).
Reverse Phase Chromatography
• It uses a non polar stationary phase and a polar mobile phase.
• Polar compounds elute faster than non polor compounds.
• When we increase polarity of mobile phase elution time will decrease.
• It Can reused / reproducible
• Mobile phase are polor compounds such as water, acetonitrile, methanol
Advantages & Disadvantages of column chromatography
Advantages
It can be used in both analytical and preparative applications.
It is used to identify the number of components of a mixture.
It is also used to separate and purify important quantities of those
components for subsequent evaluation
Any type of mixture can be separated
Any quantity of mixture can be separated
There is wider choice of Mobile Phase (Solvents)
It is low cost process and disposability of the stationary phase once it is used
in the processProcess can be scale up form lab scale
to commercial scale
Automation is possible
Disadvantages
Time consuming Process
More amounts of Mobile Phase (Solvents) required
Scale up process will take a long time to properly prepare & use
Automation makes the techniques more complicated & expensive
Types of Company we need to focus
• Pharmaceutical Industries – Bulk Drugs & API
• Nutraceuticals
• Herbal Extracts products manufacturers
• Research Laboratories
• Laboratories Chemical Repackers
• Contract Research Laboratories
Types of Department we need to contact
• R & D – Research & Development
I. Organic Synthesis Lab,
II. Medicinal Chemistry lab,
III. Novel Drug Discovery,
IV. Clinical Research,
V. Pilot Scale lab,
VI. Preparative Lab,
VII. Semi Preparative Lab
• Q.A. / Q.C.
• Process department / Production Department
• Purchase – At Last
Silent Features Manufacturing since 1973 – Consistence supplies
The product offered is highly active material
Our products has got higher surface area
The product is having better & controlled pore volumes
The Pore diameter is strictly between 50-60A
The bulk density is lower, thus you require less qty of material on column.
The product does not offer hydrolysis of your drugs after separations.
We offer batch to batch reproducible results.
Selectivity & kinetics are maintained constant ( better values)
Higher theoretical plates counts.
Manufactured under strict GMP norms
ISO 9001 accredited manufacturing firm
Comparison Between Silica Gel & Alumina Oxide
Silica Gel Alumina Oxide
Chemical For mula SiO2 (Silicon Dioxide).
Chemical Formula Al2O3 (Aluminium Trioxide ).
it is acidic in nature which we are making it to neutral
It can be Acidic, Basic & Neutral
Silica Gel has Higher Surface area as compare to Alumina i.e. 350-550 m2/gm
Alumina’s Surface Area is less than Silica Gel i.e. 140-160 m2/gm
Sizes available: 35-70, 60-120, 70-230, 100-200, 230-400 mesh
Size Available: 100-300 mesh & 200-400 mesh
Bulk Density: 040-0.65 gm/ml Bulk Density: 0.90-1.2 gm/ml
Pore Dia: 20, 60, 100, 300, 1000A Pore Dia: 50-60A
Recommended