HI Sudhakar,

First of all you should plot a graph in terms of peak area and different percentage concentration of ethanol (you can prepare the different molar concentration of ethanol and water). on comparing the area of your sample with this graph you can calculate percentage of ethanol present in your sample.

Set up a calibration curve, using standards around your predicted values - remember maximum concentration will be ~0.51g EtOH/g sugar equivalent and many times you will get much less. Mind you, EtOH density is lower then water 0.789g/cm^3 (~1ml) so if you prepare your standards by volume you have to correct for this.

Inject your standards, triplicates, and do the peak area vs. EtOH concentration. I suggest doing a pre-extraction with EthylAcetate to avoide injecting water (and even worse, sugar solution) into your GC. Another options is headspace. If you do go with extraction make sure to add an internal standard such as Iso-propanol.

See link, we have used this method on other GCs such as HP GC 5890.

Yoram

Article A simple rapid gas-chromatography flame-ionization- detector...

Determine Ethanol concentration (Ethanol Titre) as explained by Prof. Yoram Gerchman in %(v/v). Let it be X %.

Determine Sugar concentration by a suitable method in terms of g/L.

Initial Sugar concentration = So

Final Sugar concentration = Sf

Sugar Consumed = S = So - Sf

Ethanol Yield, Y = 0.789*X*10 / S    

Unit of Y = gram ethanol / gram sugar

For glucose, theoretical max value of Y = 0.51

Venkatesh Kamath is right but this will calculate ethanol Yield (i.e. how much sugar was converted to ethanol), a more 'advanced' answer then concetration per-se.

I will take this oportunity to suggest you take a look at the attached protocol (also uploaded to researchgate), more detailed then the original paper.

Yoram

Sir Yoram Gerchman you have mentioned that "I suggest doing a pre-extraction with EthylAcetate to avoide injecting water " can you explain this in detail why? Yes it is true that you are using FID with........column but we are using TCD with porapackQ column and if the temperature of oven , injector and detector is 180 then water and ethanol will be in vapor phase and so we didn't ever find any issues isn't it? Please suggest if you had ever face any issue?

GC columns In general do not react well with water and even less so with the sugar common in the fermentation broth (tend to clog the column/injector). I suggest you either do an extraction as described in the following file, if you GC is properly equiped, do Head Space (need special sampler).

Ok sir this means water is not an issue the problem is sugar and other organic compounds present in fermentation broth. I am not using to detect ethanol obtained from fermentation and so i have not ever faced such issues...Thank you.

Although not as bad as sugar, water is still an issue. Silica based column deteurate by water and it is usualy adviced not to inject water directly. See Q&A number 5 in this link

http://www.restek.com/Pages/faq_gcc#gcc5

Given your specific coulmn (i.e. porapack Q) I think that water should be less of an issue.

Yoram 

Dear,

hydrolysis 1g dried biomass with 1ml cellulase in 25ml citrate buffer --> 0.45g glucose/g biomass. Subsequently, fermentation by saccharomycei 11h-->analyze ethanol by GC. The GC results show 39%. How to calculate ethanol (g/L) or (g/g gluocose)

Thank you very much

What do you mean by "The GC results show 39%"? How did you get this number?

Yoram

the result based on ethanol standard and the amount of others compounds

OK, and how was the standart curve made? If you already know its 35% then what is the question?

I do not have experience in GC, last time I did not make the standard curve, So Must I made the standard curve? last time I only inject ethanol 99.9% and then inject sample, and the result show 39%.

My question is must I made the standard cuve? and How to calculate ethanol yield (g/L) or g/g glucose from the percentage in GC (39%)?

(hydrolysis 1g dried biomass with 1ml cellulase in 25ml citrate buffer --> 0.45g glucose/g biomass. Subsequently, fermentation by saccharomycei 11h-->analyze ethanol by GC. 

Thank you

Thuyen

To begin with GC output can't be looked upon like a pH meter or light meter output. There is no simple single output number. What you usually get is a series of peaks separated in time, hopefully one of them will be ethanol.

Your output would be peak area (and probably hight). The only "%" I can think of is % of sum of peak areas (or hight) but there is no way to convert this directly to amount of analyte as this sum will depend on many things.

What you need is a way to convert peak area to amount of analyte, i.e. a calibration curve. The most simple approach is to make a series of ethanol solutions and inject them under the expect same conditions. If you are extracting your ethanol you should also use an internal standard.

I suggest you read a little about chromatography before continuing working.

Also see my protocol in the link

Data EtOH GC-FID Protocol

Thank you very much, Gerchman,

I will follow your protocol,

may I ask you, suppose that, my sample has 0.38% ethanol (result from GC, w/w), how to calculate ethanol by g/g glucose or g/L

Thank you very much

I am not sure what w/w mean in this case (most people don't weight the water, just the EtOH) but see Venkatesh Kamath answer in the previous page and keep in mind 0.51 mass conversion is the theoretical maximum.

To explain the last point:

The reaction is (molar ratio ... 1 glucose --> 2CO2 + 2EtOH

Glucose MW = 180

CO2 MW = 44

EtOH MW = 46

So (46*2)/180=0.51

Also keep in mind EtOH density is lower then water (789g/L) so if you use a volume/volume (v/v) calibration curve you need to correct for this.

Yoram

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