Hello viewers, today we are going to check the rancidity of an oil or fat sample. To check the rancidity, we need to prepare reagents followed by conducting the crease test. Then we can determine the peroxide value and acid value and interpret the result.
To set the experiment, we need UV spectrophotometer, quartz cuvette, balance machine, incubator, falcon tube, beaker, pipette, filler, measuring cylinder, ethanol, hydrochloric acid, acetic acid, chloroform, diethyl ether, fluoro-glucanol, and trichloroacetic acid. Let's start the analysis. At first, we will prepare reagents for crease test.
30% trichloroacetic acid is needed for the quantitative test of RENCDD by Chris method. To prepare it, first measure 50 ml glacial acetic acid and take into a reagent bottle. Now take weight of 15 gm trichloroacetic acid. Transfer the trichloroacetic acid into the same reagent bottle carefully. Attach the cap and shake the bottle until the trichloroacetic acid crystal is dissolved completely.
30% trichloroacetic acid solution is now ready to use. 1% fluoro-glucanol solution is also needed for the quantitative test of RENCDT by Chris method. For the preparation of 1% fluoro-glucanol solution, measure 50 ml glacial acetic acid and take into another reagent bottle.
Take weight of 0.5 gram fluoroglucinol powder. Transfer the fluoroglucinol powder into the same reagent bottle. Shake the bottle until the fluoroglucinol powder is dissolved completely. 1% Fluoroglucinyl solution is now ready to use.
0.1% Fluoroglucinyl solution is also needed for the qualitative test of RENCDT by CRIS method. Dissolve 10 mg Fluoroglucinyl powder into 10 ml diethyl ether and shake it to dissolve the powder completely. Finally, we are ready for the CRIS test. At first, we are going for the qualitative test.
Bring the oil or fat sample and a clean and dry beaker for the sample preparation. Take 5 ml of oil or fat sample into the beaker. Add 5 ml of 0.1% protoglucinal solution into the beaker and shake it to mix the content properly. Now add 5 ml of concentrated hydrochloric acid into the beaker.
Shake the beaker to mix the content properly. Reddish pink color formation indicates the incipient rancidity. A negative control is prepared from non-rancid oil which did not develop any sort of pink color. Let's conduct the quantitative test. Bring a clean and dry falcon tube and the oil sample for sample preparation.
Take 5 ml of oil sample into the falcon tube. Add 5 level of chloroform into the tube. Close the tube with the cap and shake it to mix the content properly.
Add 10 ml of 30% trichloroacetic acid solution into the tube. Now add 1 ml of 1% fluoro-glycinal solution into the tube. Close the tube and shake well.
Incubate the test tube at 45 degrees Celsius for 15 minutes. Take out the tube after incubation. Add 4 ml of absolute ethanol into the tube.
Close the tube with the cap and rotate it gently to mix the content. Sample preparation is done. Now take the prepared sample immediately to measure the absorbance. Set the spectrophotometer wavelength at 545 nm.
Now insert the clean blank cuvette into the spectrophotometer. Click on zero icon to make the blank cuvette absorbance zero. Fill the cuvette with the prepared sample and insert the sample filled cuvette into the spectrophotometer. Click on measure icon to take the absorbance of the prepared sample.
We have got that the absorbance of the prepared sample is 0.338 which is higher than 0.2 and indicates the incipient rancidity of oil. This is a negative control which is prepared from non-branched oil. Fill the cuvette with the prepared negative control oil solution and take the absorbance as done for the sample.
We have got that the absorbance of the negative control is 0.126 which is lower than 0.15 and indicates that it is a non-rancid oil. Another way to check the RENCDT of an oil sample is the determination of peroxide value. Video link for the peroxide value determination is given in the description section.
Free fatty acid and acid value also indicate the RENCDT. Video link for the free fatty acid and acid value determination is given in the description section. Let's interpret the results.
In the qualitative CRIS test result, we saw the pink color is formed which indicates the incipient rancidity. In the quantitative CRIS test result, we got the absorbance of 0.338 which is higher than 0.2. So, we can say that the tested oil sample is a rancid oil.
Fresh oils usually have peroxide values well below 10 mEq per kg, and a rancid test often begins to be noticeable when the peroxide value is above 20 mEq per kg. Higher free fatty acid and acid values than the reference values indicate the rancidity of an oil sample.