Research SOP: STP FOR DETERMINATION OF SODIUM BENZOATE AND POTASSIUM SORBATE BY HPTLC

1.0 PURPOSE

The purpose of this document is to carry out the method of sample preparation for sodium benzoate and potassium sorbate by HPTLC.

2.0 SCOPE

This procedure is the method of sample preparation for sodium benzoate and potassium sorbate by HPTLC.

3.0 RESPONSIBILITY

The expert and trained chemist are responsible for following this procedure. The technical manager is responsible for implementing this procedure.

4.0 REQUIREMENTS

4.1 Apparatus

4.1.1 HPTLC system consisting of TLC Scanner IV, Linomat V, twin trough development chamber, and WinCats 1.4.9 evaluation software

4.1.2 TLC plate precoated with Silica gel 60 F254 (Mfg. by E. Merck, Cat. No. 5554).

4.1.3 Analytical Balance

4.1.4 Ultra sonicator

4.1.5 Mixer Grinder

4.1.6 Shaker

4.1.7 Whatman No.1 filter paper

4.2 Glassware

4.2.1 Beakers -100 ml

4.2.2 Pipette-10 ml

4.2.3 Measuring cylinder-100 ml

4.2.4 Conical flask-250 ml

4.2.5 Volumetric flask-1 Litre

4.3 Reagent

4.3.1 Sodium Benzoate

4.3.2 Potassium Sorbate

4.3.3 Chloroform

4.3.4 HPLC grade water

4.3.5 Methanol

5.0 PROCEDURE

5.1 Sample preparation
5.1.1 For Juice sample: Measure accurately about 5 ml of the test sample (properly shaken) in a 100 ml measuring cylinder. Make up the volume to 100 ml with water. Shake for 5 minutes & filter the sample with Whatman No.1 filter paper. Discard the initial 1-2 ml and then collect the filtrate and use this for HPLC analysis

5.1.2 For the Murabba sample: Take the appropriate quantity (around 100 g) of the test sample in a 1:1 ratio for the syrup to solidify and homogenize properly in the mixer grinder. Weigh accurately about 10 g of sample in a 100 ml conical flask and add 50 ml of water. Shake for 8-10 mins with intermittent shaking to disperse the sample properly in water. Filter the sample with Whatman No.1 filter paper. Discard the initial 2-3 ml and then collect the filtrate and use this for HPTLC analysis.

5.1.3 For Pickle/Chutney sample: Take the appropriate quantity (around 100 g) of the test sample and homogenized sample in a 100 ml conical flask and add 50 ml of the water. Shake for 8-10 mins with intermittent shaking to disperse the sample properly in water. Filter the sample with Whatman No.1 filter paper. Discard the initial 2-3 ml and then collect the filtrate and use this for HPTLC analysis.

5.2 Standard preparation

5.3 Dissolve 100 mg of sodium benzoate & Potassium sorbate standard separately in 100 ml water. Take 5 ml from both in a 100 ml volumetric flask and make up the volume to 100 ml of HPLC grade water.

5.4 Apply 3 µl for juice samples & 10ul for Murabba, Pickle & Chutney samples along with 2,4,6,8 µl of the standard on to TLC plate Develop the TLC plate in a solvent system consisting of Chloroform: Methanol (90:10) and allow it to run up to 90 mm.

5.5 Remove the TLC plate from the chamber and dry using a hair drier.

5.6 Scan the plate at 235 nm in absorbance mode.

5.7 Note down the area of sodium benzoate and potassium sorbate and calculate the result.

6.0 CALCULATION

By using the below equation.

y=mx +c

Where,

y=Analyte area

x= Concentration of Analyte

m=Slope of the calibration curve

c= y-axis intercept value

7.0 REVISION/CHANGE HISTORY: NA

8.0 ATTACHMENTS

Appendix-1: FLOW CHART FOR DETERMINATION OF SODIUM BENZOATE AND POTASSIUM SORBATE BY HPTLC

9.0 REFERENCES

· In-house