Method development and validation of Bronopol (Preservative) and its Stability Indicating Study by UV Spectrophotometry

 

Dr. Pankaj Kapupara,  Dhara Parekh*, Dr. Ketan Shah

Department of Quality Assurance, School Of Pharmacy, RK University, Kasturbadham, Rajkot -360 020 Gujarat, India

.*Corresponding Author E-mail: Pankaj.kapupara@rku.ac.in

 

ABSTRACT:

Preservatives are those substances that are added into pharmaceutical formulations to increase its shelf life. Prolonged use of formaldehyde in the formulations may lead to cancer and other health related issues. Besides formaldehyde has a bad odor which results in poor patient compliance. In such cases the use of bronopol can prove to be a useful option for industries. It degrades to formaldehyde but slowly and in very low amount that gives preservative action. A simple, precise and sensitive method for bronopol estimation in UV spectrophotometer was developed and validated. The method showed linearity in range of 20-100 µg/ml. LOD and LOQ values were found to be 1.22 and 3.77 µg/ml. Correlation coefficient was found to be 0.994. The developed method was also successfully applied to the pharmaceutical formulation. Stability indicating study of bronopol was also done in various conditions.

 

KEYWORDS: Bronopol . Preservative, UV Spectrophotometry, Stability indicating study.

 

 


INTRODUCTION:

“Preservatives are substances that commonly added to various foods and pharmaceutical products in order to prolong their shelf life.” They are added in formulations containing higher water content. This is done in order to avoid any alteration of composition of formulation and to avoid microbial attack that can be faced during the storage [1-2]. Formaldehyde is a preservative which is used but it is carcinogenic in nature. Formaldehyde has a bad odour which results in poor patient compliance [3]. Bronopol is an anti microbial preservative and is added to various pharmaceutical formulations to avoid their degradation. In such cases the use of bronopol can prove to be a useful option for industries. The effective concentration of bronopol in pharmaceutical preparations ranges from 0.01 to 0.1%, with the common concentration being 0.02% [4]. It degrades to formaldehyde but slowly and in very low amount that gives preservative action.

 

MATERIALS AND METHODS:

A double beam UV/Visible spectrophotometer (Labtronic-LT2900) was employed with spectral bandwidth of 1 nm and wavelength accuracy of ± 0.3 nm with automatic wavelength correction with a pair of 10 mm quartz cells. A Shimadzu electronic analytical balance (BL – 220H) was used for weighing the sample.

 

Preparation of Standard Stock Solution

Accurately weigh 100mg of standard pure dried powder of Bronopol. It was transferred into 100ml volumetric flask. Make up the volume till mark with distilled water. Final concentration of 1000µg/ml was obtained which was stock solution.

 

Selection of Analytical wavelength5

After carrying out appropriate dilution from stock solution a solution of 60µg/ml was prepared. It was scanned in the wavelength range of 200-400nm in spectrum mode. The wavelength at which absorption maxima is obtained was selected as analytical wavelength.

 

 

Selection of Analytical Range 

Appropriate quantities of volume were pipetted out from standard stock solution of Bronopol and serial dilutions were prepared. The volume was made upto the mark with distilled water to obtain concentration of 20,40,60,80 and 100 µg/ml Bronopol. The absorbances of these solutions were measured at 214nm i.e. the selected analytical wavelength. The absorbance was plotted against concentration to obtain standard calibration curve. The concentration range obeying Lambert-Beer’s law was chosen.

 

Procedure for Linearity

For preparing the solutions to determine linearity sufficient aliquots were pipetted out from standard stock solution. They were transferred to 10ml volumetric flasks. Volume was made up with the help of distilled water. Concentration range of 20, 40, 60, 80 and 100 µg/ml were prepared. Absorbance of these solutions was measured at 214nm. Absorbance Vs Concentration was plotted in order to obtain standard calibration curve. The concentration range should obey Lambert-Beer’s law.

 

Procedure for Precision

In intraday precision absorbance of standard solution containing bronopol (60 µg/mL) was measured 6 times at 214nm during the same day. %RSD was calculated. Interday precision was obtained by the assay of six sample sets on different days.

 

Stability indicating study of Bronopol

Stability study of bronopol was done in various conditions. Bronopol was kept in acidic, basic, photochemical and in hot air oven for particular period of time.

 

Acidic media :

Weigh 100 mg pure dried Bronopol powder and add 100 ml of 0.1 N HCl solution. From the above solution 1ml was withdrawn after 1, 3 and 5 hours. 1 ml of sample was taken and neutralized with 0.1N NaOH. Serial dilutions were performed in order to obtain 50μg/mL concentrated solution.

 

Basic media:

Weigh 100 mg pure dried Bronopol powder and add 100 ml of 0.1 N NaOH solution. From the above solution 1ml was withdrawn after 1, 3 and 5 hours. 1 ml of sample was taken and neutralized with 0.1 N HCl. Serial dilutions were performed in order to obtain 50μg/mL concentrated solution.

 

Photometric degradation:

Weigh 1gm pure dried Bronopol powder and put in to UV light chamber. Wavelength was kept in range 250-400nm. From this, 100 mg of sample was taken at 1, 3 and 5 hour. Serial dilutions are performed in order to obtain 50μg/mL concentrated solution.

Degradation in oven:

Weighted amount of 1 gm drug was put in to oven at 50°C temperature. From that sample after specific interval of time 1, 3 and 5 hour 100 mg of sample taken. Add in to 10 ml volumetric flask . Serial dilutions are performed in order to obtain 50μg/mL concentrated solution.

 

RESULTS:

 

 
 


Figure 1 UV Spectrum of Bronopol at 214nm

 

Table 1: Calibration table of bronopol

S.No

Concentration of Bronopol (µg/mL)

Absorbance

1

20

0.186

  2

40

0.472

 3

60

0.720

4

80

0.923

5

100

1.130

 

Table 3: LOD and LOQ for Bronopol

Drug

LOD (µg/mL)

LOQ (µg/mL)

Bronopol

1.22

3.70

 

Table 4: Intraday precision data for Bronopol

Sr. No.

Label Claim (µg/mL)

Absorbance

Label Claim (%)

1

60

0.72

100.41

2

60

0.71

99.30

3

60

0.72

100.83

4

60

0.73

101.38

5

60

0.72

100.41

6

60

0.73

101.38

 

Table 5: Inter day precision data for Bronopol

Sr. No.

Label Claim (µg/mL)

Absorbance

Label Claim (%)

1

60

0.72

100.55

2

60

0.73

101.66

3

60

0.72

100.83

4

60

0.72

100.55

5

60

0.73

102.08

6

60

0.74

103.05

There was significant degradation of bronopol found in basic conditions and when exposed to UV light. However no degradation was observed in acidic and thermal conditions.


Table 6: Statistical validation of Intraday precision data for Bronopol

Drug

Mean Label Claim (%)

Standard Deviation

Relative Standard Deviation (%)

Standard Error

Bronopol

100.62

0.77

0.77

0.31

 

Table 7: Statistical validation of Inter day precision data for Bronopol

Drug

Mean Label Claim (%)

Standard Deviation

Relative Standard Deviation (%)

Standard Error

Bronopol

101.45

1.00

0.96

0.40

 

Table 8: Stability study data of bronopol

Degradation conditions

Time (hours)

Concentration (µg/mL)

Absorbance

Amount retain (µg/mL)

Amount

Degraded (µg/mL)

Degradation (%)

NaOH

1

50

0.491

46.51

3.49

6.98

3

50

0.467

44.40

5.60

11.20

5

50

0.384

36.80

13.20

26.40

HCl

1

50

0.527

49.80

0.20

0.40

3

50

0.525

49.65

0.35

0.70

5

50

0.524

49.55

0.45

0.90

Oven

1

50

0.527

49.85

0.15

0.30

3

50

0.526

49.75

0.25

0.50

5

50

0.525

49.60

0.40

0.80

UV

1

50

0.524

49.55

0.45

0.90

3

50

0.512

48.45

1.55

3.10

5

50

0.488

46.25

3.75

7.50

 


 

Figure 2: Graphic overlay depicting Linearity over range of concentration of bronopol

 

 

Figure 3: Standard Calibration curve of Bronopol

 

DISCUSSION

Proposed method for Bronopol determination in UV spectroscopy was found to be simple, precise and sensitive. Validation data for precision are given in table 4 and 5. Stability study data for bronopol are given in table 8. The standard deviation (S.D.), relative standard deviation (%R.S.D.) and standard error (S.E.) calculated are low, indicating high degree of precision of the method. %RSD was found to be less than 2 which comply with ICH and USP. Bronopol degrades significantly in basic conditions and in UV light. Bronopol was found to be stable in acidic conditions.

 

CONCLUSION:

A simple, precise and sensitive method for bronopol estimation in UV spectrophotometer was developed and validated. This could also be applied to bronopol estimation in marketed formulation. It was also revealed through the stability indicating study of bronopol UV spectrophotometer that degradation of bronopol occurs in basic conditions and UV light. There is no significant effect of acidic media and temperature on the degradation of bronopol. So it is advisable that formulation containing bronopol should be made in acidic pH and labelled as ‘store in dark place’.

 

REFERENCES:

1.       Fahelelbom MS, Shabraway YE. Analysis of preservatives in pharmaceutical products. Pharmaceutical Reviews Year. 5(1); 2007: 5-10.

2.       Hang YH, Lai YC, Chiu CW et al. Comparing micellar electrokinetic chromatography and microemulsion electrokinetic chromatography for the analysis of preservative in pharmaceutical and cosmetic products. Journal Of Chromatography.41(5); 2003: 153-164.

3.       Rahman MM, Rashid OH, Bhadra S, Rouf AS. Development and validation of colorimetric and RP-HPLC methods for the determination of formaldehyde in cosmetics. American Chemical Science Journal. 5(3); 2015: 224-237.

4.       Lian HZ, Zhang WB et al. A study on the stability of bronopol in bronopol lotion by ion-paired reversed-phase high performance liquid chromatgraphy. Journal of Pharmaceutical and Biomedical Analysis.15; 1996: 667-671.

5.       Limbasiya AB, Kapupara PP, Shah KV. Development and Validation of Analytical Method for Simultaneous Estimation of Diclofenac potassium and Serratiopeptidase in Pharmaceutical Formulation Research J. Pharm. and Tech. 7(6); 2014:655-659

 

 

 

 

Received on 10.06.2016          Modified on 22.06.2016

Accepted on 05.07.2016        © RJPT All right reserved

Research J. Pharm. and Tech. 2016; 9(7):916-918.

DOI: 10.5958/0974-360X.2016.00175.X