Method Development and Validation of Indomethacin in Bulk Drug and Capsule Formulation by using Mix Hydrotrophy

 

Swati B. Rathod*, Poonam A. Salunke, Vaishali C. Kulkarni, Bhavika R. Chavhan, Dr. Shashikant D. Barhate

Shree Sureshdada Jain Institute of Pharmaceutical Education and Research, Jamner–424206.

 

ABSTRACT:

New, simple and rapid method indicating UV–Visible spectroscopy was developed and validated for the estimation of Indomethacin in pure form, and in formulation. Mix hydrotropic solution was used as a solvent. The adequate drug solubility and maximum assay sensitivity (320 nm) in the wavelength range of (200-400 nm), the linear calibration curve was obeyed in the concentration range of (10-50 µg/ml) show regression equation (Y=0.015X-0.002), and correlation Coefficient (R²=0.998). This method was validated and applied to the determination of Indomethacin capsules form, no interference was found from capsule excipients at the selected wavelength and analysis conditions. It was concluded that the developed method is accurate, sensitive, precise, and reproducible and indirectly for the estimation of Indomethacin content in pharmaceutical formulation.

 

 

 

INTRODUCTION:

Indomethacin is a non steroidal, anti-inflammatory agent with antipyretic, analgesic properties and is an indole  derivative designated chemically as 1-(p-chlorobenzoyl)-5-methoxy-2-methyl-1H-indole-3-acetic acid. Indomethacin is an odorless, pale yellow to yellow tan crystalline substance. It is lipid-soluble, practically insoluble in water and sparingly soluble in alcohol. Indomethacin has a pKa of 4.5 and is stable in neutral or slightly acidic media and decomposes in strong alkali. The suspension has a pH of 4.0–5.0 and it has a melting point between 155°C and 161°C and has molecular weight of 357.8. Indomethacin has a molecular formula of C₁₈H₁₆ClN0₄.

 

 

Fig no.1-Structure of Indomethacin

 

Literature survey reveals that indomethacin study was carried out by UV (hydrotrophy, KOH) ^{[3, 4, 5 ]}. RP-HPLC ^[6,7,8,9] and we developed new UV visible spectrophotometric method using Mix Hydrotropic solution as a solvent.

 

EXPERIMENTAL:

Instrumentation

UV experimentation was performed on Shimadzu 1800 UV-visible spectrophotometer equipped with Photo Diode Array (PDA) detector, with 1 cm quartz cell. Citizen Digital Ultrasonic Cleaner was used for solubility purpose.

 

MATERIALS:

All reagents used are AR grade and gift sample was obtained from Balaji drug Ahemadabad and were used as reference standard. The capsule formulation was purchased from local market.

 

Preliminary solubility study:

Solubility of drug was determined at 27±1°C. Indomethacin (10 mg) was added in 10 ml volumetric flask and 4ml of Mix hydrotropic solution (5% Sodium acetate, 5% Sodium citrate 10%, Urea) was added in it and diluted up to 10 ml with water. The clear solution of indomethacin was obtained.

 

Preparation of standard stock solutions:

Indomethacin standard stock solution: (1000 µg/ml).

An accurately weighed indomethacin (10 mg) was added in 10 ml volumetric flask and dissolved in 4ml of Mix hydrotropic solution (5% Sodium acetate, 5% Sodium citrate 10% Urea) was added in it and diluted up to 10ml with water to get final concentration (1000 µg/ml).

 

Study of spectra and selection of wavelength:

The aliquot portions of standard stock solutions of indomethacin were diluted appropriately with distilled water to obtain concentration 10 mg/ml of drug. The solutions of drug was scanned in the range of 400–200 nm. The UV absorbance spectrum of indomethacin is shown in Fig.2 From the spectrum wavelength selected for estimation of drug was 320 nm as λ max of indomethacin.

 

 

 

 

Fig.2 UV-Absorbance spectrum of indomethacin at 320 nm

 

 

Study of linearity curves:                                                                                                                                   

The aliquot portion of standard stock solutions of indomethacin was diluted appropriately with distilled water to get a series of concentration from 10-50 mg/ml for drug. The absorbance of this drug was measured at 320 nm respectively and calibration curve was plotted as concentration versus absorbance.

 

Linearity and range:

The suitable aliquots were taken to obtain 10, 20, 30, 40, 50 µg/ml from indomethacin stock solution. The results are shown in Figure no.3. indomethacin obeyed linearity in the concentration range of 10-50 μg/ml respectively at λ max 320 nm with correlation coefficient (R²>0.998) in this case. Marketed brand of capsule was analyzed.

 

 

Fig.3 Linearity curve of indomethacin at 320 nm

 

Analysis of Marketed Formulation by Proposed Method:

20 Tablets was accurately weighed, and reduced to fine powder. An accurately weighed powder sample equivalent to 10 mg of indomethacin was transferred to 10 ml volumetric flask  and  4 ml Mix hydrotropic solution (5% Sodium acetate, 5% Sodium citrate 10% Urea) was added in it and diluted up to 10 ml with water. Sonicate it for 20 min. The solution was filtered through Whatmann filter paper no. 41. The filtrate was further diluted with water to get final concentration (1000 µg/ml). From this solution 10 µg/ml was prepared. The absorbance of sample solution was measured at 320 nm and the results are shown in Table No. 1.

 

Table No. 1: Results of Application of Proposed Method for Analysis of Marketed Formulation

Sample	Label Claimed	%Label Claim*±SD	% RSD
Indocap S.R	INDOMETHACIN 75mg	100 ±0.284	0.284

*mean of each 3 reading

 

 

Validation of Method: ^[10,11]

Accuracy:

Accuracy of each of the proposed method was ascertained on the basis of recovery studies performed by standard addition method as shown in the table no.2

 

Precision:

Precision of the analytical method is expressed as the series of the measurement. It was ascertained by replicate estimation of the drug by the proposed method as shown in table no.3

 

Repeatability:

Repeatability was carrying out analysis for no. of times. The results are shown in table no. 04

LOD and LOQ:

The LOD and LOQ were estimated from the set of 5 calibration curves LOD=3.3*σ/S and LOQ=10*σ/s. LOD and LOQ was found to be 0.338 and 1.027.

 

Ruggedness:

The evaluation of Ruggedness should be considered during the development phase and depends on the type of procedure under study. It shows the reliability of an analysis with respect to external factors in method parameters such as analyst variation in table no.5.

 

 

 

 

 

Table No.2: Accuracy

Sr. no.	Level (%)	Amt. taken (µg/ml)	Amt. Added (µg/ml)	Absorbance Mean*±S.D	Amt. recovered Mean *±S.D.	%Recovery Mean*±S.D.
1	80	30	24	0.721±0.0004	24.20±0.04	100.93±0.16
2	100	30	30	0.903±0.005	30.33±0.46	101.11±1.54
3	120	30	36	1.080±0.0004	36.16±0.04	100.43±0.11

*mean of each 3 reading

 

Table No.3: Precision

Conc.(µg/ml)	Inter day			Intra Day
	Absorbance Mean±S.D.	Amt. Found	%Amt. Found	Absorbance Mean±S.D.	Amt. Found	%Amt. Found
20	0.30±0.006	20.15	100.76	0.303±0.006	20.29	101.44
30	0.45±0.001	30.20	100.67	0.452±0.001	30.24	100.81
40	0.61±0.010	40.80	100	0.603±0.006	40.33	100.83

*mean of each 3 reading

 

Table No.4: Repeatability

Sr. No.	Conc. (µg/ml)	Absorbance	Amt. Found	% Amt. Found
1	30	0.451	30.20	100.67
2	30	0.451	30.20	100.67
3	30	0.451	30.20	100.67
4	30	0.451	30.20	100.67
5	30	0.451	30.20	100.67
6	30	0.451	30.20	100.67
7	30	0.451	30.20	100.67
8	30	0.451	30.20	100.67
9	30	0.452	30.27	100.89
10	30	0.453	30.33	101.11
	Mean	0.451	30.22	100.73
	S.D.	0.0006	0.042	0.14
	%RSD	0.91	0.141	0.14

Table No.5: Ruggedness

Analyst I			Analyst II
Sr.No.	Conc. (µg/ml)	Absorbance	Sr.No.	Conc.(µg/ml)	Absorbance
1	30	0.45	1	30	0.451
2	30	0.45	2	30	0.45
3	30	0.45	3	30	0.45
4	30	0.45	4	30	0.45
5	30	0.45	5	30	0.451
6	30	0.44	6	30	0.44
	Mean	0.448		Mean	0.448
	SD	0.003		SD	0.003
	% RSD	0.831		% RSD	0.869

 

 

Label Claim:

Table no: 6. Label claim

Conc. (µg/ml)	Absorbance	Amt found	% Label Claim
30	0.451	30.2	100.66
30	0.451	30.2	100.66
30	0.449	30.06	100.222
30	0.448	30	100
30	0.447	29.93	99.77
30	0.448	30	100.00
30	0.448	30	100.00
30	0.448	30	100
30	0.448	30	100.00
30	0.448	30	100
Mean	0.4486	30.04	100
SD	0.001	0.085	0.284
%RSD	0.285	0.284	0.284

 

RESULT:

Table no: 7.

Sr.No	Validation parameter	Result
1	Solubility	Mix hydrotropic solution ( 5% sodium acetate, 5% sodium citrate, 10% urea)
2	UV detection maximum wavelength	320 nm
3	Linearity range	10-50 µg/ml
4	Standard regression equation	Y=0.015x-0.002
5	Coefficient of regression(R2)	0.998
6	%Recovery (n=9)	0.002
7	Precision Interday(n=3) Intraday (n=3)	0.001 0.001
8	Repeatability (n=6)	0.14
9	LOD LOQ	0.338 1.027
10	Ruggedeness (n=6)	0.003

 

 

DISCUSSION:

The developed method was rapid, simple, accurate for validation of indomethacin in bulk and capsule formulation. UV scanning at 400-200 nm of indomethacin shows that 320 nm is the suitable wavelength for detection of drug. (fig.2). Indomethacin shows linearity in the concentration range of 10-50 µg/ml (R²=0.998) respectively. Recovery study of drug were carried out for the accuracy parameter this study were carried out at three levels (80%, 100%, 120%) by standard addition method. The mean recovery was found to be 100.82% for indomethacin respectively. The developed method was found to be simple, precise, specific, and accurate Therefore this method can be applied for routine analysis of drugs in formulation and in bulk drug.

 

ACKNOWLEDGEMENT:

The authors are Special thankful Dr. R. K. Maheswari for their valuable gauidance and thankful to Balaji drugs for providing gift samples of indomethacin respectively.

CONCLUSION:

Indomethacin is water insoluble drug made soluble using suitable hydrotropic agent, without corrosive organic solvent and it is environment safe. The developed method is accurate, precise, robust and rapid and economical nature. Therefore this method can be applied for routine analysis of drugs in formulation and bulk drug.

 

REFERENCES:

1.       Indian Pharmacopoeia, 2007 volume 2^nd, 609.

2.       Rathod SB. Spectroscopic quantitation and analytical validation of indomethacin in bulk and capsule formulation’’ Journal of pharmacy research, 2017, 11 (2), 124-127.

3.       Jain AK. Solubilization of indomethacin using hydrotropes for aqueous injection.” European Journal of Pharmaceutics and Biopharmaceutics. 2008, 68 (3), 701-714.

4.       Ali KF. New assay method UV spectroscopy for determination of indomethacin in pharmaceutical formulation ’’Journal of Chemical and Pharmaceutical Research. 2005, 7 (4), 1591-1596.

5.       Maheshwari R.K., Rathore A., “New spectrophotometric estimation of indomethacin capsules with niacinamide as hydrotropic solubilizing agent” Pharmaceutical method 2011, 2 (3), 184-188.

6.       Kougioumtzoglou A., peikova, m. georgieva, al. zlatkov. Evaluation of the stability of indomethacin substance under a model of physiological conditions, using modified and validated RP _HPLC method”. Pharmacia, 2015, 62 (2), 10-17.

7.       Sawant N. Development and validation of a stability indicating RP-HPLC method for the estimation of indomethacin in pharmaceutical capsule formulation” Journal of Pharmaceutical Research. 2016, 15 (3), 79-85.

8.       Nandy BC. Gupta AK. Mazumder B. Development and validation of analytical methods of indomethacin in rabbit blood plasma by RP-HPLC technique. Journal of Biomedical and Pharmaceutical Research.2013, 2 (5), 01-06.

9.       Virginia B. Beverley G. and Alan N. High-Performance liquid chromatographic assay of indomethacin in porcine plasma with applicability to human levels. Journal of chromatographical Science. 2006, 44 (1), 41-44.

10.     Potdar MA. Pharmaceutical Quality Assurance, 3^rd Ed. Nirali Prakashan, Pune, 2013, 8.28-8.31.

11.     Mohamed Ahmed. Hydrotropic and cosolvent solubilisation of indomethacin’’ Acta pharm. 49 (1999), 291-298.

 

 

 

 

Received on 09.05.2018        Modified on 10.07.2018

Accepted on 16.07.2018     ©A&V Publications All right reserved