I need to analyze total sugar level in my sample (common food item, especially ready to eat food). I don’t wish use complex method like HPLC. Hence, I used following procedure. I used CRM (corn based baby food from FAPAS), but the recovery values is not good (nearly 180%). So, do you have any comments for below method or do you have any suitable method?
Procedure:
Standard solution prepared using Glucose -stock solution
Sample Preparation
Weigh 2 g from the sample interested
Transfer it into 200 mL beaker & add nearly 100 mL distilled water
Boil the sample for 10-15 minutes while stirring in a water bath
Add 3 mL of conc. H2SO4 into the sample and again boil for 3 mins.
Filter the sample into 250 mL volumetric flask using fluted filter paper
Sample dilution
Take 2.5 mL from the sample into 100 mL volumetric flask and volume up to to the mark
Again take 5 mL from above sample into 50 mL conical flask and add 15 mL conc. H2SO4, 5 mL 5% phenol as previously done.
Measure the absorbance with respect to the color development at 492 nm using UV-Visible spectrophotometer.
Total CHO % in sample = (sample concentration *dilution factor) / 10000
Dear Bedigama,
Please see attached the original paper which describes phenol-sulphuric acid method. You may need to revise the quantity of reagents that you are using. If you decide to use 5% phenol solution the reatios of sample to phenol solution to sulphuric acid should be 1:1:5. Also, suggested wavelength is 490 nm. To calculate the percentage of total carbohydrates you may need to use the equation as follows: ((reading from the standard curve in micrograms/1 000 000 (to express your result in grams)/(sample weight x dilution factor)) x100%. I hope this helps.
Have you looked into this sugar meter?
http://hannainst.com/usa/prods2.cfm?id=039001&ProdCode=HI%2096801
and this
http://www.linearinstrument.com/digital-refractometers.html
The best method would be by HPLC, which will give you details of the sugars which then can be summed. Colorimetric methods will err on the low side, since some some di- or trisaccharides are not measured until laborious premeasurement hydrolysis.
You can also use the anthrone method. carbohydrates are hydrolyzed with dilute hydrochloric acid to simple sugars. The anthrone reagent is used to develop a green colour . Absorbance which is proportional to the developed colour complex is read at 620 or 630nm in a UV-visible spectrophotometer. a standard such as glucose is used to calibrate and quantify the amount of sugar in the sample. You can check this reference. Hedge, J E and Hofreiter, B T (1962) In: Carbohydrate Chemistry 17 (Eds Whistler R L and Be Miller, J N) Academic Press New York
The sugars may be extracted in 95% ethanol, evaporated at 80 ° C and subjected to acid hydrolysis with concentrated HCI, boiling for 15 minutes, according & Lane Eynon (1934). These can be quantified by the Somogyi method (Nelson, 1944) by reduction of cupric to cuprous ion, which reacts with the arsenic-reactive molibídico forming molybdenum oxide, whose concentration is proportional to the amount of sugars in the sample.
I have noticed that you have a very long name. What should I call you?
There is a simple method for total sugar content. This method is Dubois et al. (1956) using a standard for example glucose.
If you have no time or possibility , you can sum up glucose, fructose and saccharose contents after HPLC analysis in plants. Or check this ms
Article Analytical methods for determination of sugars and sweetness...
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