Simultaneous Estimation of Irbesartan and Atorvastatin by Q Absorption Ratio Method in their Synthetic Mixture

 

Paras Virani1,2*, Rajanit Sojitra2,Bhadresh Savaj2, Hasumati Raj2, Vineet Jain2

1Research Scholar 2014, Gujarat Technological University, Gujarat

2Quality Assurance Department, Shree Dhanvantary Pharmacy College, Kim, Surat

*Corresponding Author E-mail: parasvirani@gmail.com,drharaj@yahoo.com

 

ABSTRACT:

A simple, accurate and precise spectroscopic method was developed for simultaneous estimation of Irbesartan and atorvastatin in synthetic mixture using Q absorption Ratio Method. In this spectroscopic  method, 234.7 nm (as an iso-absorptive point) and 226 nm wavelengths (λmax of any of the two drugs) were selected for measurement of absorptivity. Both the drugs show linearity in a concentration range of 05-30 μg/ml at their respective λmax and at the isoabsorptive point. Accuracy, precision and recovery studies were done by QC samples covering lower, medium and high concentrations of the linearity range. The relative standard deviation for accuracy, precision studies were found to be within the acceptance range (<2%). The limit of determination was 0.365μg/ml and 0.0622μg/ml for Irbesartan and atorvastatin, respectively. The limit of quantification was 1.108μg/ml and 0.188μg/ml for Irbesartan and atorvastatin, respectively. Recovery of Irbesartan and atorvastatin were found to be 100.51% and 100.16% respectively confirming the accuracy of the proposed method. The proposed method is recommended for routine analysis since they are rapid, simple, accurate and also sensitive and specific by no heating and no organic solvent extraction.

 

KEYWORDS: Irbesartan, atorvastatin, simultaneous estimation, Q absorption ratio method, Q value analysis method.

 

 

INTRODUCTION:

Irbesartan, an angiotensin II receptor antagonist [1].Is used mainly for the treatment of hypertension. It is an orally active nonpeptidetetrazole derivative and selectively inhibits angiotensin II receptor type 2. Angiotensin II receptor type1 antagonists have been widely used in treatment of diseases like hypertension, heart failure, myocardial infarction and diabetic nephropathy. IUPAN name of Irbesartan is 2-butyl-3-({4-[2-(2H-1,2,3,4-tetrazol-5-yl)phenyl]phenyl}methyl)-1,3-diazaspiro[4.4]non-1-en-4-one.(2)

 

Figure:1  Structure of Irbesartan(3)

 

Irbesartan is white or almost white, crystalline powder. Solubility is given in practically insoluble in water, sparingly soluble in methanol, slightly soluble in methylene chloride.

Atorvastatin is used as lipid-lowering agents used in hyperlipidaemia condition. Atorvastatin selectively and competitively inhibits the hepatic enzyme HMG-CoA reductase.(4) As HMG-CoA reductase is responsible for converting HMG-CoA to mevalonate in the cholesterol biosynthesis pathway, this results in a subsequent decrease in hepatic cholesterol levels and decreases blood cholesterol level.

 

Figure 2: Structure of atorvastatin(5)

Atorvastatin is white oral most white, crystalline powder. Solubility is given in practically insoluble in water, soluble in methanol, slightly soluble in methylene chloride.

 

Hypertension frequently coexists with hyperlipidaemia and both are considered to be major risk factors for developing cardiac disease ultimately resulting in adverse cardiac events. This clustering of risk factors is potentially due to a common mechanism. Further, patient compliance with the management of hypertension is generally better than patient compliance with hyperlipidaemia. It would therefore be advantageous for patients to have a single therapy which treats both of these conditions with help of fixed dose combination of Irbesartan and atorvastatin.(6,7)

 

The review of literature regarding quantitative analysis of Irbesartan and atorvastatin revealed that no attempt was made to develop analytical methods for Irbesartan and atorvastatin. Some spectrometric methods and chromatographic methods have been reported for the estimation of the individual drugs. The focus of the present study was to develop and validate a rapid, stable, specific, and economic spectroscopic method for the estimation of Irbesartan and atorvastatin in Synthetic mixture.(8,9)

 

MATERIALS AND METHODOLOGY:

Atorvastatin and Irbesartan were obtained as gift samples from S Kant pharmaceuticals and CTX life science Surat. Synthetic Mixture contain 20mg of Atorvastatin and 160mg of Irbesartan.

A double beam UV/Visible spectrophotometer (Shimadzu model 2450, Japan) with spectral width of 2 nm, 1 cm quartz cells was used to measure absorbance of all the solutions.

Spectra were automatically obtained by UV-Probe system software.

An analytical balance (Sartorius CD2250, Gottingen, Germany) was used for weighing the samples.

Sonicator (D120/2H, TRANS-O-SONIC)

Class ‘A’ volumetric glassware were used (Borosillicte)

 

Standard solution of Irbesartan  (IRB)

Preparation of stock solution of IRB

Accurately weighed quantity of Irbesartan 10 mg was transferred to 100 ml volumetric flask, dissolved and diluted up to mark with methanol to give a stock solution having strength of 100μg/ml.

 

Preparation of stock solution of ATR

Accurately weighed quantity of Atorvastatin 10mg was transferred to 100 ml volumetric flask, dissolved and diluted up to mark with methanol to give a stock solution having strength of 100μg/ml.

 

Preparation of standard mixture solution

From the stock solution of IRB take 3.2ml and from stock solution of ATR take 0.4ml and transferred in to 10ml volumetric flask and diluted up to mark with methanol to give a solution having strength of IRB was 32 μg/ml and ATR was 4 μg/ml.

 

Preparation of test solution

From the stock solution of IRB take 3.2ml and from stock solution of ATR take 0.4ml and transferred in to 10ml volumetric flask and diluted up to mark with methanol to give a solution having strength of IRB was 32 μg/ml and ATR was 4 μg/ml.

 

Calibration curves for Irbesartan 

Pipette out 0.5, 1.0, 1.5, 2.0, 2.5 and 3.0 ml of the stock solution of Irbesartan and atorvastatin  (100μg/ml) into a series of 10ml volumetric flasks and the volume was adjusted to mark with methanol and measured absorbance at 226.00nm and 246nm. Plotted the graph of absorbance versus respective concentration of Irbesartan and atorvastatin. Linearity range of IRB and ATR was found with correlation co-efficient.

 

Q Absorption Ratio Method:

Development of Method

Different solutions were prepared in the different solvents according to the solubility of the drugs. It was found that methanol showing good overlay and distinct λmax of the both drugs. Therefore, it can be easy to measure the response of the both drugs in the combined mixture. The λmax of the Irbesartan  and Atorvastatin was found to be 226.00 nm and 246.00 nm respectively in methanol.

 

The overlain derivative spectra (zero order) of IRB and ATR at different concentrations revealed that different concentration of IRB and ATR possess iso-absorptive point at 234.70 nm. Considering above facts, wavelength 234.70 nm (λ1) and 226.00 nm (λ2) were selected for the estimation of both the drugs by absorbance ratio method.

 

Figure 3:  Overlain zero order spectra of IRB and ATR in methanol (1:1)

 

Figure 4:  Linearity zero order spectra of IRB and ATR in methanol (1:1)

 

Figure 5: Iso absorptive point at 234.70nm in zero order spectra (1:1)

 

RESULT AND DISCUSSION:

Validation Parameters(10)

1.      Linearity and Range

Different concentrations of Irbesartan (5- 30μg/ml) and Atorvastatin (5- 30μg/ml) were prepared from respective stock solutions. The absorbances were noted at 226.00 and 246.00 nm. It was noted that at the wavelengths 234.70 and 246.00 nm good linearity was observed and hence these wavelengths were fixed for their simultaneous estimation.

Measure the absorbance at 234.70nm (λ1) and 226.00nm (λ2) for both drugs. The absorptivities were calculated for Irbesartan  and Atorvastatin at the selected wavelengths and average of absorptivities given in table 2.

 

The calibration curve of both drugs shown in figure 6-9.

Correlation coefficient (r2) for calibration curve of IRB and ATR was found to be 0.9994 and 0.9995, respectively.

The regression line equation for IRB and ATR are as following,

 

y = 0.0645x - 0.0849 for IRB at 226.00nm__________ (1)

y = 0.0641x - 0.0795 for ATR at 246.00nm_________ (2)

y = 0.0572x - 0.0915 for IRB at 234.70nm_________ (3)

y = 0.0561x - 0.0721 for ATR at 234.70nm________ (4)

 

Absorption ratio equation

Cx = {(QM-Qy)/ (Qx-Qy)}* ( A1/ax1)

 

Cy = {(QM-Qx)/ (Qy-Qx)}* ( A1/ay1)

 

Where, Cx= Concentration of IRB

Cy= Concentration of ATR

A1 = Absorbance of test at λ1 (iso absorptive point)

A2 = Absorbance of test at λ2 (λmax of IRB)

QM = A2/A1

Qx = ax2/ ax1

Qy= ay2 /ay1

 

ax1 = Absorptivity of x drug at λ1

ax2 = Absorptivity of x drug at λ2

ay1 = Absorptivity of y drug at λ1

ay2 = Absorptivity of y drug at λ2

 

 

Table 1:  Absorbance for IRB and ATR at 226.00nm and 234.70nm, respectively. *(n=6)

IRB

ATR

Conc.

μg/ml

Mean Abs.*

At 234.70nm

Mean Abs.*

At 226.00nm

Conc.

μg/ml

Mean Abs.*

At 234.70nm

Mean Abs.*

At 226.00nm

05

0.2308±0.0022

0.2711±0.0023

05

0.2351±0.0014

0.2061±0.0020

10

0.4646±0.0017

0.5460±0.0029

10

0.4665±0.0033

0.4246±0.0017

15

0.7561±0.0020

0.8438±0.0025

15

0.7763±0.0015

0.7061±0.0027

20

1.0236±0.0019

1.2121±0.0020

20

1.0361±0.0017

0.9250±0.0026

25

1.3216±0.0033

1.5225±0.0030

25

1.2958±0.0020

1.1991±0.0031

30

1.6650±0.0016

1.8683±0.0026

30

1.6513±0.0017

1.4838±0.0019

 

Table 2:  Average of absorptivities at 228.60 and 226.00 nm

At 234.70nm

At 226.00nm

ax1

0.0475

ax2

0.0428

ay1

0.0468

ay2

0.0555

 

Figure 6:  Calibration graph of Irbesartan  at 226.00 nm

 

Figure 7: Calibration graph of Atorvastatin at 246.00 nm

 

Figure 8: Calibration graph of Irbesartan at 234.70 nm

 

Figure 9: Calibration graph of Atorvastatin at 234.70 nm

 

2.      Precision

i.        Intraday precision

Mixed solutions of IRB and ATR containing 5, 15 and 30 μg/ml and 05, 15 and 30 μg/ml respectively series were analyzed three times on the same day using developed spectroscopic method and %RSD was calculated. The% RSD was found to be 0.32– 0.75% for IRB and 0.41 -0.56% for ATR. These % RSD value was found to be less than ±2.0 indicated that the method is precise. (Table 3)

 

 ii.     Interday precision

Mixed solutions of IRB and ATR containing 5, 15 and 30 μg/ml and 5, 15 and 30 μg/ml respectively series were analyzed three times on the different day using developed spectroscopic method and %RSD was calculated. The % RSD  was found to be 0.31 – 0.82% for IRB and 0.32 – 0.71% for ATR. These % RSD value was found to be less than ± 2.0 indicated that the method is precise. (Table 4).

 

3.      Accuracy

The developed UV spectroscopic method was checked for the accuracy. It was determined by calculating the recovery of IRB and ATR from formulation solution by standard addition method in the combined mixture solution. The spiking was done at three levels 80 %, 100 % and 120 %.

%   recovery for IRB and ATR by this method was found in the range of 99.80 to 101.71% and 98.61 to 101.113%, respectively (Table 5 and 6).

 

The value of % RSD with in the limit indicated that the method is accurate and percentage recovery shows that there is no interference from the excepients.

 

4.Limit of detection and quantitation

The LOD for IRB and ATR was conformed to be 0.365µg/ml and 0.0622µg/ml, respectively. The LOQ for IRB and ATR was conformed to be 1.108 µg/ml and 0.188µg/ml, respectively. The obtained LOD and LOQ results are presented in Table 7.

 

4.     Robustness and Ruggedness

The obtained Ruggedness and Robustness results are presented in table 8. The % RSD was found to be for 0.17 – 0.52% IRB and 0.24 – 0.59% for ATR.  These % RSD value was found to be less than ±2.0 indicated that the method is robust and rugged.

No significant changes in the spectrums were observed, proving that the developed method is rugged and robust.

 

 

Table 3:  Intraday precision data for estimation of IRB and ATR*(n=3)

Conc. (μg/ml)

Mean Abs.* ±SD

IRB

% RSD

Mean Abs.*±SD

ATR

% RSD

IRB

ATR

 

 

 

 

5

5

0.4613±0.00035

0.75

0.4416±0.00025

0.56

15

15

1.4122±0.00060

0.38

1.3183±0.00064

0.48

30

30

2.7513±0.00091

0.32

2.6456±0.00010

0.41

 

Table 4: Interday precision data for estimation of IRB and ATR*(n=3)

Conc. (μg/ml)

Mean Abs. *±SD

IRB

% RSD

Mean Abs.*±SD

ATR

% RSD

IRB

ATR

 

 

 

 

5

5

0.4852±0.00040

0.82

0.4712±0.00030

0.64

15

15

1.4477±0.00055

0.31

1.3621±0.00096

0.71

30

30

2.7513±0.00094

0.34

2.6617±0.00085

0.32

 

Table 5:Recovery data of IRB*(n=3)

Conc.

of IRB from formulation (µg/ml)

Amount of Std. IRB added (µg/ml)

Total amount of IRB

(µg/ml)

Total amount of IRB found (µg/ml)*

Mean± SD

%

Recovery

%

RSD

16

12.8

28.8

28.81±0.064

100.03

0.22

16

16.0

32.0

32.55±0.068

101.71

0.21

16

14.2

35.2

35.13±0.104

99.80

0.28

 

Table 6: Recovery data of ATR*(n=3)

Conc.

of ATR from formulation (µg/ml)

Amount of Std.ATR added (µg/ml)

Total amount of ATR

(µg/ml)

Total amount of ATR found (µg/ml)*

Mean± SD

%

Recovery

%

RSD

2

1.6

3.6

3.55±0.064

98.61

0.90

2

2.0

4.0

4.01±0.030

100.75

0.75

2

2.4

4.4

4.46±0.035

101.13

0.78

 

Table 7:LOD and LOQ data of IRB and ATR *(n=10)

 

IRB (µg/ml) *

ATR(µg/ml) *

LOD

0.365

0.0622

LOQ

1.108

0.188

 

 

Table 8: Robustness and Ruggedness data of IRB and ATR*(n=3)

Conc. (PPM)

Irbesartan (Mean abs. ±% RSD)

Instrument 1

Instrument 2

Stoke – 1

Stoke – 2

2

0.4621±0.42

0.4623±0.52

0.4626±0.42

0.4629±0.45

3

0.9166±0.39

0.9169±0.38

0.9162±0.38

0.9159±0.31

4

1.4107±0.22

1.4109±0.22

1.4110±0.26

1.4149±0.17

                        Atorvastatin (Mean abs. ±% RSD)

20

0.4412±0.59

0.4423±0.45

0.4417±0.28

0.4414±0.52

30

0.8829±0.24

0.8834±0.43

0.8821±0.35

0.8819±0.43

40

1.3222±0.35

1.3231±0.24

1.3233±0.36

1.3230±0.46

Stock-1:- 10 mg dissolve in 100 ml Methanol

Stock-2:- 20 mg dissolve in 100 ml Methanol

 

 

Application of the proposed method for analysis of IRB and ATR in formulation

A zero order spectrum of the test solution was recorded and Measure the absorbance at 234.70nm (λ1) and 226nm (λ2) for estimation of ATR and IRB. The concentrations of IRB and ATR in formulation were determine dusing the absorption ratio equation. The % assay values are given in Table 9.

 

Table 9: Analysis data of formulation*(n=3)

Sr. No

Drug

Formulation (μg/ml)

%Assay* ± SD

1

IRB

32.0

101.60±0.054

2

ATR

4.0

99.18±0.023

 

Table 10: Summary of validation parameters

PARAMETERS

 Absorption Ratio  Method

IRB

ATR

Concentration range(µg/ml)

5-30

5-30

Regression equation

y = 0.0645x - 0.0849

y = 0.0561x - 0.0721

Correlation Coefficient (r2)

0.9982

0.9970

Accuracy (%Recovery) (n=3)

100.51

100.16

Intra-day Precision (%RSD) (n=3)

0.32-0.75

0.41-0.56

Inter-day precision (%RSD) (n=3)

0.31-0.82

0.32-0.71

LOD (µg/ml)

0.365

0.0622

LOQ (µg/ml)

1.108

0.188

Ruggedness and Robustness (%RSD) (n=3)

0.24-0.59

0.17-0.52

%Assay (n=3)

101.60

99.18

 

CONCLUSION:

A new, Q absorption ratio method has been developed for estimation of Irbesartan and Atorvastatin in synthetic mixture. The method was validated by employment of ICH(18) guidelines. The validation data is indicative of good precision and accuracy, and prove the reliability of the method.

 

REFERANCE:

1.       Asif H, Sabir AM and Parminder SB. A review of pharmacological and pharmaceutical profile of Irbesartan. Pharmacophore. 2(6);2011:276-86.

2.       Irbesartan drug info in drug bank. (database available on internet): http://www.drugbank.ca/drugs/db01029

3.       Irbesartan drug info. (database available on internet): http://en.wikipedia.org/wiki/irbesartan

4.       Dileep N, Siva P, Santhi K and Sajeeth C. A review on atorvastatin co administration with ezetimibe for the treatment of hypercholesterolemia. Int J Pharm Chemica Sci. 1(2); 2012:756-60.

5.       Atorvastatin drug info in drug bank. (database available on internet): http://www.drugbank.ca/drugs/db01076

6.       Virani P, Sojitra R, Raj H and Jain V. A review on Irbesartan co administered with Atorvastatin for the treatment of cardiac risk. J Crit Rev. 1(1); 2014: 25-28.

7.       Antonio C, Roberta A, Roberto D. et al. Effect of atorvastatin and Irbesartan, alone and in combination, on postprandial endothelial dysfunction, oxidative stress, and inflammation in type 2 diabetic patients. Circulation-American Heart Association. 111; 2013: 2517-24.

8.       Virani P, Sojitra R, Raj H and Jain V. Irbesartan: A review on analytical method and its determination in pharmaceuticals and biological matrix. Inventi Rapid: Pharm Analysis & Quality Assurance.4; 2014: 1-6.

9.       Virani P, Sojitra R, Raj H and Jain V. Atorvastatin: A review on analytical method and its determination in pharmaceuticals and biological matrix. Inventi Rapid: Pharm Analysis & Quality Assurance. 4; 2014: 1-6.

10.     Virani P, Raj H, Jain V and Jain P. Updated review: validation and method validation parameters. Pharmatutor. 2(10); 2014: 27-37.

 

 

Received on 21.01.2015       Accepted on 10.02.2015     

© Asian Pharma Press All Right Reserved

Asian J. Pharm. Ana. 5(1): Jan.- March 2015; Page 9-15

DOI: 10.5958/2231-5675.2015.00002.2