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Eurasian Journal of Analytical Chemistry ISSN: 1306-3057

2017 12(3):211-222 DOI 10.12973/ejac.2017.00164a

© Authors. Terms and conditions of Creative Commons Attribution 4.0 International (CC BY 4.0) apply.

Correspondence: Umang H. Shah, Ramanbhai Patel College of Pharmacy, Charotar University of Science and

Technology, India.

[email protected]

Chemometric Assisted Spectrophotometric Methods for Simultaneous Determination of Paracetamol and

Tolperisone Hydrochloride in Pharmaceutical Dosage Form

Umang H. Shah Charotar University of Science and Technology, INDIA

Ankita H. Jasani Charotar University of Science and Technology, INDIA

Received 19 June 2016 ▪ Revised 6 September 2016 ▪ Accepted 6 September 2016

ABSTRACT

Three simple Thermometric assisted UV- Visible Spectrophotometric methods, Classical

Least Square (CLS), Partial Least Square (PLS) and Principal Component Regression (PCR)

were developed for simultaneous estimation of PCM and TOL in pharmaceutical dosage

form without any chemical separation and any graphical treatment of the overlapping

spectra of two drugs. The UV absorption spectra of the drugs studied in the range of 220 –

280 nm. Beer’s law was obeyed for both drugs in the concentration ranges of 5 – 25 µg/ml

for Paracetamol and 1.5 – 7.5 µg/ml for Tolperisone Hydrochloride. Twenty five (25) mixed

solutions were prepared for the chemo metric calibration as training set and sixteen mixed

solutions were prepared as validation set. The absorbency data matrix was obtained by

measuring the absorbency at twenty-one wavelength points, from 220 to 280nm with the

interval of 3nm. The developed methods can be applied in simultaneous determination of

the selected drugs from the pharmaceutical formulation in routine analysis.

Keywords: PCR, chemometric, paracetamol (PCM), tolperisone hydrochloride (TOL), PLS

and CLS

INTRODUCTION

Paracetamol (PCM) chemically it is, 4-hydroxyacetanilide [Figure 1(a)] is a well-known

analgesic drug. It is used for temporary relief of fever, minor aches and pains. It is official in

Indian Pharmacopoeia (IP), European Pharmacopoeia (EP), Japanese Pharmacopoeia (JP),

British Pharmacopoeia (BP) and United State Pharmacopoeia (USP) [1-5]. Tolperisone

Hydrochloride (TOL) chemically, it is (2R, S)-2-Methyl-1-(4-methylphenyl)-3-piperidin-1-

propan-1-one mono hydrochloride [Figure 1(b)] is a piperidine derivative. TOL is official in

Japanese Pharmacopoeia (JP) [6]. It is a centrally acting muscle relaxant. The combination of

PCM and TOL is used for the treatment of adult patients with acute muscle/musculoskeletal

spasms. The combination of PCM and TOL is commercially available in tablet dosage form.

mailto:[email protected]

U. H. Shah & A. H. Jasani

212

Literature review revels that numbers of analytical methods like RP-HPLC [7-9] and

UV Spectrophotometry [10-14] are available for determination of PCM and TOL in

combination and individual dosage forms and combination with other drugs .The reported

UV spectrophotometric methods are based on multicomponent analytical methods viz.

simultaneous equation, absorbance ratio and first derivative methods. These methods are

based on the univariate or bivariate calibration methods.

In the present work, the chemometric (multivariate calibration methods) approach was

applied for the multicomponent analysis of drug substances with a spectrophotometric

method. Multivariate calibration is a chemometrics method which has been employed for

determination of drugs in combined dosage [15-19], this study aims to introduce an alternative

analytical procedure based on the chemometric-assisted spectrophotometric methods for the

analysis of PCM and TOL in tablet dosage form and three multivariate calibration methods

like principle component regression (PCR), partial least square regression (PLS) and classical

least square (CLS) were developed and validated for the determination of PCM and TOL

combination in tablet dosage form.

EXPERIMENTAL

Materials and reagents

Reference standard of PCM and TOL were procured as gratis sample from Alembic

Pharmaceuticals Ltd. (Baroda, Gujarat, India) and Zydus Cadila Healthcare Ltd. (Ahmedabad,

Gujarat) respectively. Distilled water was prepared using Millipore SmartPak® DQ5

Purification Pack system.

Instrumentation and software

Shimadzu AUX 220, electronic balance was used for weighing the samples. The

Shimadzu UV-1800, UV-Visible double beam spectrophotometer with a matching pair of 1 cm

quartz cuvettes (Shimadzu Corporation, Kyoto, Japan), connected to a computer loaded with

Shimadzu UVPC version 3.42 software was used to record UV spectra of solutions. The

spectral band width was 0.5 nm. Unscrambler® 10.3.0.80, MATLAB (R2009A) 7.8.0.347 and

Microsoft excel were used for PCR, PLS and CLS model development and data analysis.

HO

HN CH3

OH3C

CH3

O

N

.HCl

(a)(b)

Figure 1. (a) Chemical Structure of PCM, (b) Chemical Structure of TOL

Eurasian J Anal Chem

213

Preparation of standard stock solution

Accurately weighed and transferred 10 mg of PCM and TOL working standard into two

different 100 mL volumetric flask respectively, and volume was made up to 100 mL with

distilled water. The final concentration of PCM and TOL were 100 µg/mL of each.

Preparation of working stock solution

Standard stock solution of PCM and TOL were used as a working solution.

Table 1. Composition of calibration set data

Sr. No. PCM (µg/ml) TOL (µg/ml) Sr. No. PCM (µg/ml) TOL (µg/ml)

1 15 4.5 14 15 7.5

2 15 1.5 15 25 7.5

3 5 1.5 16 25 1.5

4 5 7.5 17 5 6

5 25 3 18 20 1.5

6 10 7.5 19 5 4.5

7 25 4.5 20 15 6

8 15 3 21 20 6

9 10 3 22 20 3

10 10 6 23 10 1.5

11 20 7.5 24 5 3

12 25 6 25 10 4.5

13 20 4.5

Table 2. Composition of validation set data

Sr. No. PCM (µg/ml) TOL (µg/ml)

1 8 2.4

2 8 3.9

3 13 3.9

4 13 6.9

5 23 3.9

6 13 2.4

7 8 5.4

8 18 5.4

9 18 3.9

10 13 5.4

11 18 2.4

12 8 6.9

13 23 6.9

14 23 5.4

15 18 6.9

16 23 2.4


214

Construction of Calibration set

The samples for calibration set were prepared in distilled water by using mutually

orthogonal design. Total 25 mixtures were prepared for calibration set which are shown in

Table 1. Absorbance of UV spectrums was recorded in the wavelength range 220-280 nm at 3

nm wavelength interval.

Construction of Validation set

The samples for validation set were prepared in distilled water. Total 16 mixtures were

prepared for validation set which are shown in Table 2. Absorbance of UV spectrums was

recorded in the wavelength range 220-280 nm at 3 nm wavelength interval.

RESULTS AND DISCUSSION

Calibration matrix and selection of spectral zones for analysis by CLS, PCR and

PLS

Figure 2 has shown the overlain zero-order spectra for PCM and TOL individually and

in mixed in distilled water. As shown in figure PCM exhibit absorption maxima at 243 nm and

TOL exhibit absorption maxima at 261 nm. Two Chemometric calibrations, using the zero-

order spectra, were separately applied to simultaneous determination of these drugs in

mixtures. The absorbance matrix was obtained by measuring the zero-order absorbance in the

wavelength range between 220 and 280 nm, as shown. The quality of multicomponent analysis

is dependent on the wavelength range and the spectral mode used. Original and reconstructed

spectra of the calibration matrix were compared in order to select the range of wavelengths

along with using the root mean squared error of cross validation (RMSECV) and root mean

squared error of prediction (RMSEP) values. The wavelength range 220 - 280 nm with 3 nm

intervals was selected, since this range was providing the greatest amount of information

about the two components.

Figure 2. Overlay Spectra of PCM, TOL and Mixture


215

Statistical analysis in selecting the number of principal components or factors

The ability of a calibration can be define in several ways. In this sub-section, we were

calculated the standard variation of chemometric calibrations in the case of investigated

mixtures. An appropriate choice of the number of principal components or factors is necessary

for PCR and PLS calibrations. A cross-validation method leaving out one sample at a time was

employed. With a calibration set of 25 calibration spectra, PLS, CLS and PCR calibrations on

24 calibration spectra were performed, and using this calibration the concentration of the

sample left out during the calibration process was determined. This process was repeated 25

times until each calibration sample had been left once. The predicted concentrations were

compared with the known concentrations of the compounds in each calibration sample.

To validate the model, both RMSECV and RMSEP were considered; they must be as low

as possible for a particular model. RMSECV and RMSEP were calculated for each method as

per equation 1 and 2.

Table 3. Recovery studies of PCM and TOL by PCR method

Expected

Conc. (µg/ml)

Predicted Conc.

(µg/ml)

% Recovery Residual Conc.

(Expected -

Predicted) (µg/ml)

(Expected -

Predicted)2 Conc.

(µg/ml)

PCM TOL PCM TOL PCM TOL PCM TOL PCM TOL

8 2.4 8.011 2.427 100.14 101.13 -0.011 -0.027 0.000 0.001

8 3.9 8.028 3.961 100.34 101.57 -0.028 -0.061 0.001 0.004

13 3.9 13.146 3.911 101.12 100.27 -0.146 -0.011 0.021 0.000

13 6.9 13.070 7.044 100.54 102.09 -0.070 -0.144 0.005 0.021

23 3.9 22.733 3.909 98.84 100.23 0.267 -0.009 0.071 0.000

13 2.4 12.974 2.467 99.80 102.78 0.026 -0.067 0.001 0.004

8 5.4 8.030 5.514 100.37 102.11 -0.030 -0.114 0.001 0.013

18 5.4 17.960 5.451 99.78 100.95 0.040 -0.051 0.002 0.003

18 3.9 17.991 3.949 99.95 101.25 0.010 -0.049 0.000 0.002

13 5.4 13.035 5.425 100.27 100.47 -0.035 -0.025 0.001 0.001

18 2.4 17.973 2.430 99.85 101.25 0.027 -0.030 0.001 0.001

8 6.9 8.006 7.043 100.08 102.07 -0.006 -0.143 0.000 0.020

23 6.9 22.460 6.762 97.65 98.00 0.540 0.138 0.291 0.019

23 5.4 23.110 5.493 100.48 101.71 -0.110 -0.093 0.012 0.009

18 6.9 17.976 6.951 99.86 100.74 0.024 -0.051 0.001 0.003

23 2.4 22.383 2.373 97.32 98.86 0.617 0.027 0.381 0.001

Mean % 99.77 100.97

SDa 1.017 1.231

RSDb 1.020 1.220

SEP Value 0.229 0.082

RMSEPc 0.222 0.079

(a=Standard Deviation, b=Relative Standard Deviation c= Root-Mean-Square Error of Prediction)


216

RMSECV = √∑(𝐶𝑎𝑐𝑡 − 𝐶𝑝𝑟𝑒𝑑)

2

𝐼𝑐 Equation 1

where,

RMSECV= Root Mean Square Error of Cross Validation

Cact = Actual concentration of the calibration set samples

Cpred = Predicted concentration of the calibration set samples

Ic = Total number of calibration set samples

RMSEP = √∑(𝑌𝑎𝑐𝑡 − 𝑌𝑝𝑟𝑒𝑑)

2

𝐼𝑝 Equation 2

where,

RMSEP= Root Mean Square Error of Prediction

Yact= Actual concentration of the prediction set samples

Ypred= Predicted concentration of the prediction set samples

Ip= Total number of prediction set samples.

Table 4. Recovery studies of PCM and TOL by PLS method

Expected

Conc. (µg/ml)

Predicted Conc.

(µg/ml)


(Expected -

Predicted) (µg/ml)

(Expected -

Predicted)2 Conc.

(µg/ml)


8 2.4 7.975 2.426 99.69 101.07 0.025 -0.026 0.001 0.001

8 3.9 8.013 3.960 100.16 101.53 -0.013 -0.060 0.000 0.004

13 3.9 13.145 3.910 101.12 100.25 -0.145 -0.010 0.021 0.000

13 6.9 13.058 7.043 100.45 102.07 -0.058 -0.143 0.003 0.021

23 3.9 22.775 3.907 99.02 100.17 0.225 -0.007 0.051 0.000

13 2.4 12.968 2.465 99.75 102.71 0.032 -0.065 0.001 0.004

8 5.4 8.000 5.512 100.00 102.08 0.000 -0.112 0.000 0.013

18 5.4 17.973 5.450 99.85 100.93 0.027 -0.050 0.001 0.003

18 3.9 17.966 3.947 99.81 101.20 0.034 -0.047 0.001 0.002

13 5.4 13.031 5.424 100.24 100.44 -0.031 -0.024 0.001 0.001

18 2.4 17.971 2.428 99.84 101.17 0.029 -0.028 0.001 0.001

8 6.9 7.980 7.042 99.75 102.06 0.020 -0.142 0.000 0.020

23 6.9 22.419 6.759 97.47 97.96 0.581 0.141 0.338 0.020

23 5.4 23.049 5.490 100.21 101.67 -0.049 -0.090 0.002 0.008

18 6.9 17.957 6.949 99.76 100.71 0.043 -0.049 0.002 0.002

23 2.4 22.369 2.370 97.26 98.76 0.631 0.030 0.398 0.001

Mean % 99.65 100.92

SDa 0.995 1.238

RSDb 1.000 1.230

SEP Value 0.221 0.077

RMSEPc 0.214 0.075


217

The predicted concentrations of the components in each sample were compared with the

actual concentrations of the components in each validation samples and the root mean square

error of cross validation (RMSECV) was calculated for each method. The RMSECV was used

for examining the error in the predicted concentrations. The model is a key to achieving correct

quantitation in PLS, CLS and PCR calibrations. The resulted models were also validated by

prediction of the concentration of analytes in separate validation set which was not used in the

model development. The results of prediction and the % recoveries are represented in Table 3

to 5. The predictive abilities of the models were evaluated by plotting the actual known

concentrations against the predicted concentrations which are shown in Figure 3. Figure has

shown there was good agreement between the predicted (calculated) and actual concentration

of drugs. The means recoveries and the relative standard deviation of our proposed methods

were computed and indicated in Table 3 to 5 for PCM and TOL, respectively. Another

diagnostic test was carried out by plotting the concentration residuals against the predicted

concentrations. Figure 4 has shown the residuals appear randomly distributed around zero,

indicating adequate models building. Satisfactory correlation coefficient (r2) and slope values

were obtained for each compound in the validation set by PLS, CLS and PCR optimized

models indicating good predictive abilities of the models.

Table 5. Recovery studies of PCM and TOL by CLS method

Expected Conc.

(µg/ml)

Predicted Conc.

(µg/ml)


(Expected -

Predicted) (µg/ml)

(Expected -

Predicted)2 Conc.

(µg/ml)


8 2.4 7.947 2.444 99.34 101.83 0.053 -0.044 0.003 0.002

8 3.9 7.973 3.998 99.67 102.52 0.027 -0.098 0.001 0.010

13 3.9 13.121 3.939 100.93 101.01 -0.121 -0.039 0.015 0.002

13 6.9 13.004 7.108 100.03 103.02 -0.004 -0.208 0.000 0.043

23 3.9 22.691 3.949 98.66 101.27 0.309 -0.049 0.096 0.002

13 2.4 12.937 2.487 99.51 103.61 0.064 -0.087 0.004 0.008

8 5.4 7.940 5.568 99.25 103.11 0.060 -0.168 0.004 0.028

18 5.4 17.933 5.505 99.63 101.94 0.067 -0.105 0.005 0.011

18 3.9 17.924 3.995 99.58 102.43 0.076 -0.095 0.006 0.009

13 5.4 12.977 5.478 99.82 101.45 0.023 -0.078 0.001 0.006

18 2.4 17.924 2.454 99.58 102.25 0.076 -0.054 0.006 0.003

8 6.9 7.957 7.102 99.46 102.92 0.043 -0.202 0.002 0.041

23 6.9 22.348 6.835 97.16 99.05 0.652 0.065 0.426 0.004

23 5.4 22.979 5.550 99.91 102.77 0.021 -0.150 0.000 0.022

18 6.9 17.894 7.017 99.41 101.69 0.106 -0.117 0.011 0.014

23 2.4 22.298 2.400 96.95 100.01 0.702 0.000 0.492 0.000

Mean % 99.31 101.93

SDa 0.994 1.192

RSDb 1.000 1.170

SEP Value 0.267 0.117

RMSEPc 0.259 0.113


218

Figure 3. (a) PCR – Expected vs. Predicted Concentration of PCM, (b) PCR – Expected vs. Predicted

Concentration of TOL, (c) PLS – Expected Vs. Predicted Concentration of PCM, (d) PLS – Expected Vs.

Predicted Concentration of TOL, (e) CLS – Expected vs. Predicted Concentration of PCM, (f) CLS – Expected

vs. Predicted Concentration of TOL

Table 6. Assay results of PCM and TOL by developed PCR, PLS and CLS methods (n=6)

Drug Amount

Taken

(µg/ml)

Amount Found (µg/ml) Mean % ± SDa % RSDb

PCR PLS CLS PCR PLS CLS PCR PLS CLS

PCM 15 15.107 15.014 15.196 100.31

±

0.910

100.72

±

0.832

100.97

±

0.308

0.907 0.826 0.305

15.049 14.971 15.168

15.107 15.027 15.073

15.236 15.123 15.186

14.863 15.242 15.122

14.918 15.268 15.131

TOL 4.5 4.584 4.570 4.576 101.49

±

0.277

101.49

±

0.286

101.71

± 0.221

0.273 0.282 0.217

4.575 4.562 4.578

4.564 4.580 4.585

4.573 4.549 4.560

4.555 4.582 4.574

4.552 4.558 4.589

a=Standard Deviation, b=Relative Standard Deviation)


219

Assay of Marketed formulation

Twenty tablets were accurately weighed and finely powdered. Tablets powder

equivalent to about 500 mg of PCM and 150 mg of TOL accurately weighed and transferred to

into 100 mL amber colored volumetric flask and 70 mL of distilled water was added. The

mixture was sonicated for 20 min and diluted up to the mark with distilled water and filtered

through a whatman filter paper no.41. From this solution 1 mL aliquot was withdrawn into a

100 mL amber colored volumetric flask and diluted up to the mark with water to get the

solution containing 50 μg/mL of PCM and 15 μg/mL of TOL. From this solution 3 mL aliquot

was withdrawn into a 10 mL amber colored volumetric flask and diluted up to mark with

water. So solution contains 15 μg/mL of PCM and 4.5 μg/mL of TOL. The analysis procedure

was repeated six times for tablet formulation. The result was shown in Table 6.

Figure 4. (a) PCR – Expected vs. Residual Concentration of PCM, (b) PCR – Expected Vs. Residual Concentration of

TOL, (c) PLS – Expected Vs. Residual Concentration of PCM, (d) PLS – Expected Vs. Residual Concentration of TOL,

(e) CLS – Expected Vs. Residual Concentration of PCM, (f) CLS – Expected Vs. Residual Concentration of TOL


220

Accuracy Study

The accuracy of the method was carried out at three levels 80 %, 100 % and 120 % of the

working concentration of the sample. The calculated amount of a standard solution of PCM

and TOL was spiked with added sample solution to prepare level 80 %, 100 % and 120 % of

the working concentration. The analysis procedure was repeated for three times. The result

was shown in Table 7 and 8.

SUMMARY AND CONCLUSION

Three chemometric methods were applied successfully to simultaneous determination

of PCM and TOL in pharmaceutical dosage form. The summary parameters of all three

chemometric methods were shown in Table 9. Model that gave lowest RMSECV values when

used for predicting the unknown samples, predicted well by giving lowest RMSEP values. On

the other hand, the fundamental advantages of investigated methods are the simultaneous

analysis of the mixture of the subject drugs, without chemical pre-treatment, speed of analysis

Table 7. Accuracy data of PCM by PCR, PLS and CLS methods

Level % Amount

Taken

(µg/ml)

Amount Found (µg/ml) Mean % ± SDa % RSDb


80 % 18 18.262 18.221 18.250 101.24

±

0.198

100.99

±

0.252

100.30

±

0.246

0.205 0.251 0.245

18.195 18.195 18.189

18.226 18.138 18.276

100 % 20 20.147 20.200 20.117 100.11

±

0.596

100.06

±

0.853

100.22

±

0.913

0.605 0.852 0.914

19.919 19.871 19.845

20.024 19.965 20.182

120 % 22 21.853 21.952 21.807 99.44

±

0.974

99.74

±

0.457

99.51

±

0.801

0.988 0.463 0.813

22.107 22.032 22.096

21.685 21.837 21.779

(a=Standard Deviation, b=Relative Standard Deviation)

Table 8. Accuracy data of TOL by PCR, PLS and CLS methods

Level % Amount

Taken

(µg/ml)

Amount Found

(µg/ml)

Mean % ± SDa % RSDb


80 % 5.4 5.431 5.434 5.431 100.67 ±

1.113

100.67 ±

1.113

100.62 ±

1.109

1.110 1.111 1.104

5.385 5.383 5.387

5.500 5.506 5.506

100 % 6 5.985 5.988 5.989 99.52

±

1.308

99.52

±

1.308

99.41

±

1.303

1.315 1.315 1.313

5.897 5.894 5.883

6.046 6.042 6.042

120 % 6.6 6.521 6.524 6.510 99.09

±

0.465

99.09

±

0.465

98.93

±

0.460

0.477 0.479 0.470

6.585 6.589 6.561

6.534 6.532 6.528

(a=Standard Deviation, b=Relative Standard Deviation)


221

and cost effectiveness. Hence, the developed methods can be applied in simultaneous

determination of the selected drugs from the pharmaceutical formulation in routine analysis.

ACKNOWLEDGEMENT

The author(s) thank to Alembic Pharmaceuticals Ltd. and Zydus - Cadila Healthcare Ltd. for

providing gift samples PCM and TOL and Ramanbhai Patel College of Pharmacy

(CHARUSAT), Changa for providing the necessary infrastructure to carry out the research.

REFERENCES

1. British Pharmacopoeia. (2009). British Pharmacopoeia Commission, the Medicine and Healthcare Products Regulatory Agency. London, 2, 4548.

2. European Pharmacopeia. (2011). European Directorate for the Quality of Medicines and Healthcare, 2, 2667.

3. Indian Pharmacopoeia. (2010). Govt. of India Ministry of Health and Family Welfare, New Delhi, 3, 1859.

4. United States pharmacopoeia. (2009). United States Pharmacopoeial convention, Rockville, 31, 1024. 5. Japanese Pharmacopeia. (2011). Japanese Directorate for the Quality of Medicines and Healthcare, 2,

1515. 6. Japanese Pharmacopeia. (2011). Japanese Directorate for the Quality of Medicines and Healthcare, 2,

318. 7. Gowramma, B., Rajan, S., Muralidharan, S. & Meyyanathan, S. (2010). A validated RP-HPLC

method for simultaneous Estimation of Paracetamol and Diclofenac Potassium in pharmaceutical formulation. International Journal of Chem Tech Research, 2, 676.

8. Vaghela, V., & Koladiya, B. (2012). Development and validation of a RP-HPLC method for the estimation of Tolperisone Hydrochloride in bulk and pharmaceutical dosage form. International Journal of Advances in Pharmaceutical Analysis, 2, 6.

Table 9. Summary parameters of chemometric methods

Parameters PCM TOL

PCR PLS CLS PCR PLS CLS

Range (µg/ml) 5 - 25 1.5 – 7.5

Wavelength (nm) 220 - 280 220 - 280

Δλ (nm) 3 3

Factor 7 7 - 7 7 -

% Recovery 99.77 99.65 99.31 100.97 100.92 101.93

SD 1.017 0.995 0.994 1.231 1.238 1.192

RSD 1.020 1.000 1.000 1.220 1.230 1.170

Correlation

coefficient (r2)

0.999 0.999 0.999 0.998 0.998 0.9985

Intercept 0.978 0.978 0.9758 1.0039 1.004 1.0154

Slope 0.249 0.246 0.226 - 0.015 - 0.015 - 0.0582

SEC Value 0.102 0.077 0.256 0.106 0.086 0.201

SEP Value 0.229 0.221 0.267 0.079 0.077 0.117

RMSECV 0.100 0.076 0.250 0.104 0.084 0.197

RMSEP 0.222 0.214 0.259 0.082 0.075 0.113


222

9. Sireesha, K. & Kumar, R. (2012). Simultaneous estimation of Lornoxicam and Tolperisone by RP - HPLC. International Journal of Advances in Pharmaceutical Research, 3, 981.

10. Shah, U., Kavad, M., & Raval, M. (2013). Development and validation of UV spectrophotometric method for estimation of Paracetamol and Flupirtine Maleate in bulk and pharmaceutical dosage form. International Journal of Pharm Tech Research. 5, 1007.

11. Sawant. R., Bhangale, L., Joshi, R., & Lanke, P. (2010). Validated spectrophotometric methods for simultaneous estimation of Paracetamol, Domperidone and Tramadol HCl in pure and tablet dosage form. Journal of Chemical Metrology, 4, 21.

12. Mahaparale, P., Shinde, S., & Nirmal, N. (2013). Simultaneous UV – Spectrophotometric Estimation of Diclofenac and Tolperisone Hydrochloride in tablet dosage form. International Journal of Research and Development in Pharmacy and Life Sciences, 2, 574.

13. Shah, U., Thula, K., Raval, M., & Desai, P. (2012). Development and validation of UV spectrophotometric methods for simultaneous estimation of Tolperisone Hydrochloride and Paracetamol from combined tablet dosage form. International Journal of Biological & Pharmaceutical Research, 3, 634.

14. Patel, G., Parmar, R., Nayak, P., & Shah, D. (2012). The simultaneous estimation of Paracetamol and Tolperisone Hydrochloride in tablet by UV Spectrophotometric methods. Journal of Pharmaceutical Science and Bio scientific Research, 2, 63.

15. Sankar, A., Vetrichelvan, T., & Venkappaya, D. (2011). Simultaneous estimation of Ramipril, Acetylsalicylic acid and Atorvastatin calcium by chemometrics assisted UV-spectrophotometric method in capsules. Acta Pharmaceutica, 61, 283.

16. Dinç, E., & Baleanu, D. (2002). Spectrophotometric quantitative determination of cilazapril and hydrochlorothiazide in tablets by chemometric methods. Journal of pharmaceutical and biomedical analysis, 30, 715.

17. El-Gind, A., Emara, S., & Shaaban, H. (2007). Development and validation of chemometrics-assisted spectrophotometric and liquid chromatographic methods for the simultaneous determination of two multicomponent mixtures containing bronchodilator drugs. Journal of Pharmaceutical and Biomedical Analysis, 43, 973.

18. Mohamed, A. E.-M. I., & Mikre, W. (2009). Determination of lamivudine and stavudine in pharmaceutical preparations using chemometric sassisted spectrophotometry. Saudi Pharmaceutical Journal, 17, 275.

19. Shah, U., Patel, S., Raval, M., & Desai, P. (2015). Chemometric assisted spectrophotometric methods for the simultaneous determination of Rifampicin and Piperine in bulk and capsule. Indian Journal of Pharmaceutical Education and Research, 49, 200.

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Chemometric Assisted Spectrophotometric Methods for ... Charotar University of Science and Technology, INDIA ... based on the univariate or bivariate calibration ... Gujarat, India)