I have digested a waste newspaper with mild sulphuric acid and want to estimate carbohydrate content before and after treatment.
Have you tried HPLC? you can try running your hydrolysate on HPLC using Sugar-pack column or other compatible column. here is the condition that can be used for sugar determination using Sugar-pack column: The mobile phase: deionized MilliQ water (resistivity ~ 18 Mohm) containing 5 mg/L CaNa2-EDTA. Flow rate: 0.6 mL/min. Column maintained at 80°C. Alternatively, you can use Rezex column which use dilute sulfuric acid as eluent (refer here: http://www.instrument.com.cn/Quotation/Manual/103999.pdf). You should run standard in similar condition, and therefore you can determine which peaks of samples have similar retention time as standard.
You can also try to use TLC as described in this paper: http://assbt-jsbr.org/JSBR/Vol20/JSBRVol20No3P251to254SeperationandAnalysisofSomeSugarsbyUsingThinLayerChromatography.pdf
hope this help.
You can use an anion-exchange column designed specifically for high-resolution separations of mono- and disaccharides. HPLC separation is accomplished with a simple isocratic eluent, and detection is accomplished with pulsed amperometric detection (PAD). No derivatization is required.
Best and simple way to estimate,quantitate specially hexose in your sample is to use phenol/sulfuric acid assay and read it at 450. Also, you can quantiate the amounts of all sugars by using HPAE-PAD HPLC.
Dear Ioannes Kyriakou, thanks for your response. The treatment is with sulphuric acid.
Are you aiming to get the total hexose in the sample? if yes, then phenol sulphuric acid, reducing sugar (such as Somogyi-Nelson or DNS) will work. However, if you want to determine what sugars (and also the quantity of the sugars) present, you should use HPLC to differentiate between classes. Or, if you specifically want to get the glucose concentration, you can use Glucose Oxidase assay, which specific only for glucose. Other sugar in the solution will not give interferences.
After digestion, the phenol-sulphuric acid or the anthrone method could be used to quantify the inherent carbohydrates. A standard food analysis text will give you the details of the methods. You need a standard depending on the sugar of interest. the carbohydrates would have been hydrolyzed to their component sugars. Both methods can be used to develop colors hat can be estimated spectrophotometrically at specific wavelengths. Alternatively, chromatography could be used
MALDI-MS is also a viable option, though the data would be more difficult to analyze than HPLC or LC-MS, which are likely the best options.
presence only (shiffer& smogy procedure, I think) or determination of amount?.
hydrolyzable carbohydrate or reducing and non-reducing sugars?
I think a lot of methods
yeah i want the presence and amount also. Eshak, can you help me know the shiffer and smogy procedure.
An additional pertinent consideration to extraction of hexoses from newspapers by my own thinking would be that newspapers may not contain significant amounts of hexoses considering the raw material from which newspapers are made.
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