INTRODUCTION:

Rosuvastatin is chemically [(E,3R,5S)-7-[4-(4-fluorophenyl)-2-[methyl(methylsulfonyl)amino]-6-propan-2-ylpyrimidin-5-yl]-3,5-dihydroxyhept-6-enoic acid] Calcium salt with empirical formula (C₂₂H₂₇FN₃O₆S)₂Ca. RSV is classified under class II of BCS¹.

RSV reduces Low Density Lipoproteins (LDL) levels by increasing the LDL receptors on the cell surface and internalize the circulating LDL². It acts by inhibiting 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMG-CoA reductase), the rate limiting enzyme in cholesterol biosynthesis.

The typical dose of RSV Calcium is 5-40mg per day and strongly reversibly bound to plasma protein having a prolonged effect on hepatic cholesterol synthesis in animal models².

Chromatographic techniques are time-consuming and expensive thus a simple and accurate validated UV-spectrophotometric method can be a great alternative for routine pharmaceutical research. Many solvents are hazardous in nature and have to replace with green solvents for analysis of drugs^3-5.

The FDA Inactive Ingredients Database lists sodium lauryl sulphate as a commonly considered safe excipient. One of the twelve principles of green chemistry is to use safer solvents in chemical processes to prevent pollution caused by poisonous and volatile organic solvents⁶.

Hence the main objective of the present investigation is to implement the use of green solvent in estimation of RSV. The chemical structure of RSV is given in Figure 1.

Figure 1. Structure of RSV Calcium

METHOD AND MATERIALS:

Instrumentation:

UV-Spectrophotometer of Shimadzu UV-1900 with Lab Solutions software and Shimadzu UV-1800 with UV Probe software were used for determination of RSV. Calibrated weighing balance was used for weighing.

Drug Sample:

RSV (API) was obtained as a gift sample from Apotex Research Pvt. Ltd. Bengaluru.

Reagents and Chemicals:

Sodium Lauryl Sulfate and other chemicals used for the experiment were obtained from the store house of KLE College of Pharmacy, Belagavi.

Selection of Wavelength:

RSV is soluble in various organic solvents but, to develop a safe and greener method, the green solvent 2 % Sodium Lauryl Sulfate was selected throughout the study. By using a UV-Spectrophotometer, RSV 10µg/ml of working standard solution was scanned between 400nm and 200nm, with maximal absorption at 242 nm. ⁷.

Preparation of stock solution:

In a 100ml clean and dried volumetric flask, a carefully weighed quantity of 10mg of RSV calcium was taken. To dissolve the medication, a 2 percent SLS solution was added, and the volume was increased to 100ml with the same. With a concentration of 100µg/ml, this was considered a standard stock solution. Further dilutions were made using the standard stock solution⁸.

Plotting of calibration curve:

From the standard stock solution, serial dilutions containing concentrations of 5, 10, 15, 20 and 25mg/ml were prepared. The solutions were analyzed for 5 sets and the absorbance were measured at 242nm. Calibration curve was plotted as Concentration on x-axis and Absorbance on y-axis and linear regression equation was calculated⁹.

Method development and validation:^10-20

Various solvents were used for the estimation RSV was found to be soluble and stable for 1 week at room temperature in 2% SLS solvent. Therefore, this solvent was used for the determination of suitable detection wavelength and working concentration of standard. The ICH has produced criteria for validation of analytical methods, namely Q2 (R1), which describes this procedure as characteristic performance verified by laboratory research. The UV spectrophotometric method developed was tested according to validation requirements for analytical processes in order to demonstrate the method's appropriateness utilizing optimal parameters.

Linearity was tested in the 5-25µg/ml range. Statistical calculations were used to determine the LOD and LOQ. Six duplicates of a solution containing 15µg/ml of RSV were created to test system precision, and the absorbance of each solution was measured at 242nm and the %RSD computed.

The precision of the method was evaluated by performing assays under (i) Intraday precision and (ii) Interday precision. Six duplicates of a solution containing 15µg/ml RSV were evaluated for intraday precision, and % RSD was calculated at various time intervals on the same day. Six duplicates of a solution containing 15µg/ml RSV were evaluated for Interday precision, and the % RSD was calculated across three days. Ruggedness was determined by testing the same suggested method on multiple instruments and by different analysts to ensure reproducibility, which revealed a % RSD less than 2, indicating that the presented approach is robust. Recovery experiments were used to determine accuracy, which involved determining the percent mean recovery of a sample using the percentage approach at three distinct levels. From the dilutions used for linearity, 50-150 percent of the sample solutions were produced according to the process outlined in the procedures. For the analysis RSV was taken in the concentrations of 5, 10, 15mg/ml. At each level, three analyses were performed. Percent mean recovery was calculated. The established recovery limits are 98-102%, and all observed data fits within this range, indicating good recovery values and the absence of interference by typical pharmaceutical additives and excipients. We have also performed Solution and stock solution stability. The % RSD for absorbance obtained by fresh dilutions containing RSV was calculated.

RESULTS AND DISCUSSION:

UV-spectrophotometric method was developed by using green solvent and details of method developed were presented in Table 1. Spectrum of solvent and spectrum of RSV is presented in Figure 2 and 3 respectively.

Table 1: Developed method parameters.

Sr. No.	Parameters	Specifications
1	Analytes	RSV
2	Solvent	2% Sodium Lauryl Sulfate
3	λmax	242nm

Figure 2. UV-Spectrum of Solvent

Figure 3. UV-Spectrum of RSV

Method showed linear response between the concentration range of 5-25mg/ml with r² value 0.999. 1.58µg/ml and 4.79µg/ml were found to represent the LOD and LOQ, respectively. All the %RSD values obtained for precision were found to be less than 2% and results were present in the Table no.2, Table no.3, Table no.4, Table no.5 and Table no.6. The accuracy of RSV from tablet dosage forms was found between the 97-102%.

Table 2: Linearity and range data of RSV

Sr. No.	Concentration (mg/mL)	Absorbance
1	0	0
2	5	0.152
3	10	0.363
4	15	0.559
5	20	0.741
6	25	0.966
R²=0.999
y=0.038817143

Table 3: System precision data of RSV

Replicates	Concentration (mg/ml)	Absorbance
1	15	0.552
2	15	0.556
3	15	0.561
4	15	0.566
5	15	0.566
6	15	0.568
Average		0.561
SD		0.005
% RSD		1.14%

Table 4: Intraday precision data of RSV

Replicates	Concentration (mg/ml)	Absorbance 1 Hr	Absorbance 4 Hr	Absorbance 8 Hr
1	10	0.330	0.332	0.334
2	10	0.333	0.334	0.336
3	10	0.327	0.330	0.333
4	10	0.320	0.336	0.331
5	10	0.334	0.332	0.328
6	10	0.331	0.331	0.331
Average		0.329	0.332	0.332
SD		0.005	0.002	0.002
% RSD		1.55%	0.65%	0.84%

Table 5: Interday precision data of RSV

Replicates	Concentration (mg/ml)	Absorbance 1^st Day	Absorbance 2^nd Day	Absorbance 3^rd Day
1	15	0.574	0.538	0.566
2	15	0.580	0.544	0.573
3	15	0.577	0.548	0.558
4	15	0.586	0.547	0.557
5	15	0.581	0.552	0.553
6	15	0.580	0.532	0.554
Average		0.579	0.543	0.560
SD		0.004	0.007	0.007
% RSD		0.7%	1.35%	1.39%

Table 6: Ruggedness and solution stability data of RSV

Replicates	Concentration (mg/ml)	Absorbance Analyst -1	Absorbance Analyst -2	Absorbance Solution Stability
1	15	0.336	0.332	0.328
2	15	0.332	0.331	0.326
3	15	0.331	0.333	0.329
4	15	0.337	0.336	0.331
5	15	0.334	0.336	0.336
6	15	0.330	0.332	0.334
Average		0.33	0.33	0.33
SD		0.0028	0.0022	0.0038
% RSD		0.841	0.648	1.142

Analysis of marketed formulation:

The established method for determining RSV in Tablets was used. Twenty tablets were tested, and the results showed that the amount of medicine in tablet samples was in good agreement with the formulation's label claim, with a percent assay of 98.88%.

CONCLUSION:

The developed method for estimating RSV in pharmaceutical dose form and in bulk can be concluded to be simple, sensitive, accurate, exact, reproducible, inexpensive, and environmentally friendly. The excipients in the commercial sample evaluated had no effect on the results, demonstrating the method's specificity for this formulation.

ABBREVATION:

1. UV- Ultraviolet

2. RSV- Rosuvastatin

3. SLS- Sodium Lauryl Sulphate

4. ICH- International Council for Harmonization

5. LOD- Limit Of Detection

6. LOQ- Limit Of Quantitation

7. r²- Correlation coefficient

8. RSD- Relative Standard Deviation

9. SD- Standard deviation

ACKNOWLEDGEMENT:

The authors are thankful to Principal Dr. S. S. Jalalpure and Vice Principal Dr. M. B. Patil for their support and guidance. Authors are thankful to Ms. Prathima Rajput, Mr. Tushar Powalkar and Ms. Esha Jain for helping during the research work.

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Received on 13.08.2020 Modified on 23.03.2021

Research J. Pharm.and Tech 2022; 15(2):587-590.

DOI: 10.52711/0974-360X.2022.00096