You can use spectrophotometric method using Glucose oxidase-Peroxidase kit...

I recommend you Somogyi-Nelson for reducting sugars. It's easy, quick and cheap.

Depending on the required accuracy, we sometimes use a generic device for blood sugar measurement used to control diabetes. Due to high production numbers, this device is quite cheap and gives quick results. Yet it shouid be tested with a fermentation sample to check for applicability.

You can use the enzymatic analysis kit. However, you need a glucose analyser instrument for that.

DNS or Somogy is the best and simplest for determination of reducing sugars

I think the GOD-POD reagent method is the simplest and quickest for glucose quantification. Alternatively, you can try the membrane based method on a biochemical analyzer but these membranes are only short lived.

Depends on how much accuracy are you looking for. I prefer hplc as thats the gold standard in almost all cases, but DNSA or enzyme analysis are quicker and give you agood estimate.

DNS.

Take a look.

http://www.scribd.com/doc/401070/Use-of-Dinitrosalicylic-Acid-Reagent-for-Determination-of-Reducing-Sugar

An enzyme kit is useful, generates gluconic acid, is simple and quick

I would suggest to go for DNSa method or GOD-POD by enzymes, or HPLC ... and if you have sensors in fermentors for online-estimation, then it will be fastest and online, though it is expensive technique.

I would suggest you to use GOP-POD kit that will help you to find the amount of Glucose released during fermentation, that is the easy method. DNS does not tells you the exact glucose it will shows you the total sugars

The anthrone method...

Enzymatic kit (Boheringer)

The enzymatic glucose kit (Boehringer). If you buy the reagents and enzymes individually, it is very cost effective if you have large quantities of samples to do. We also do this for glucose and fructose determination.

If you have a plate spectrophotometer and platform shaker, you can modify the method to be used in a 96 well plate (manual handling or robot handling of liquids). In 45min you could measure 88 samples with 8 standards on a single plate.

Hope this is helpful.

The fastest is a strip assay for blood glucose you can buy in a drug store

go to google, and search for ATAGO.

ATAGO products have variuos type of pen typed glucose detection meter. can read within 5 sec.

Have a try and you may double confirm it using HPLC after got all the sampling done.

Gd luck

Hi everyone am very happy with the feedback thank you very much for the help, hopefully sometime in the future i could be able to assist back.

many thanks

Sam

I have used DNS, Anthrone method for total sugar and reducing sugar estimation which roughtly takes about 45 min to 60 min. for estimation. Later switched over to glucose enzyme kit i.e. blood glucose monitor kit calibrated in the lab and it gives glucose estimation less than 1 min. and helps in calculating my glucose feed.

Using sugar analyzer (i.e; YSI analyzer equipment). I involved with these heaps glucose analysis from yeast-glucose fermentation and this machine is really user-friendly machine....all the best

Most faster way to quantify glucose will be -DNSA reducing sugar analysis.This takes 10 min and faster than GOD POD kit.

for fermentation sample DNS assay will be best and also cost reduction

Hi

DNS is a cualitative method... so if you need numbers you must to modifique the assay to get good aproximations (samples known concentration).

I would like to know how obtain good results (numerics) without HPLC.

God bless you.

Glucose sensor for blood sugar works well. 10 seconds. typical sensors are e.g CONTOUR XT from Bayer for 40 $.

I used it over years with good results - especially for starch hydrolysates. For straw hydrolysates or molasses i would not trust before calibration..

There are several methods of estimation of glucose in fermentation broth. One can use DNSA method for estimating glucose in the media, But this will be offline one. If you are looking to feed glucose to maintain certain concnetration of it in the media then this may not be the best one. In that case there are on-line glucose analyzers available and you can couple your feed rate with the analyzer and maintain a desired glucose concentration through out the batch.

The simplest test for glucose is using Keto-Diastix Reagent Strips for Urinalysis ( 2883) which is used by diabetics routinely to check the glucose level in urine. The cost is $ 5-6 for box with 50 strips.

DNS is the best method, it is fast and accurate.

Cheers evryone really appreciated...

Using a hospital glucose analyer. The sensitivity is not extremely high though. Check with YSI biochemisttry analyer

Ezymatic kit

I agree with the DNS method. However, it cannot be compared with HPLC in terms of accuracy.

If you have high glucose concentration then Brix polarimeter is good too for estimation or trend of glucose concentration during fermentation, I do not know the price of it but i think it is somewhere in range of 500-1000 $.

DNS method with pre calibrated for Glucose is the right test for laboratory methods

Two methods given below will give you an estimation of glucose, in fact both of them if done accurately would give the same answer.

DNS for reducing sugar,

Phenol sulphuric acid for total sugar

To measure glucose but not maltose, maltotriose and longer sugar oligomers, we use:

GOD-Perid glucose reagent (glucose assay)

Phosphate buffer, pH 7.5 250mM

Phenol 5mM

4-aminoantipyrine 0.5mM

Glucose oxidase (GOD) >10kU/l

Peroxidase (Perid) >1 kU/l

Deionised water

Glucose standard (1 mg/ml) for making your own dilution series

This glucose assay is a typical two stage enzymatic assay where all components are present in excess except for the primary substrate (glucose) that we want to measure. In the first step, glucose oxidase converts glucose and oxygen to glucuronic acid and hydrogen peroxide (H2O2). The released peroxide then produces quinoneimine from 4-aminoantipyrine and phenol under the catalytic action of peroxidase. Quinoneimine can then be measured spectrophotometrically. You have to make a glucose standard curve and then measure your unknown samples. Have fun. HPLC is ideal for checking non-fermentable sugars (maltotetraose, maltopentaose etc....)

Hi .....

You can go for Dinitro Salicylic Acid (DNS) Method. Regarding DNS method, reducing sugars have the property to reduce 3,5-dinitrosalicylic acid (DNS) reagent. This method tests for the presence of free carbonyl group (c=o), the so called reducing sugar. This involves the oxidation of the aldehyde functional group present;for example; in glucose and the ketone functional group in fructose. Simultaneously 3,5-DNS in alkaline solution is reduced to 3 amino 5 nitro salicylic acid. The above reaction scheme shows that one mole of sugar will react with one mole of 3,5 dinitrosalicylic acid. Different reducing sugars generally yield different colour intensities ;thus it is necessary to caliberate for each sugar. In addition to the oxidation of the carbonyl groups in the sugar, other side reactions such as the decomposition of sugar also competes for the availability of 3,5 dinitrosalicylic acid.

Next option anthrone method where Glucose is dehydrated to hydroxymethyl furfural. This compound forms with anthrone a green coloured product with an absorption maximum at 630 nm. If you are working with complex substrate like polymers, then better you select Anthrone method where it works in the presence of strong acid like H2SO4.

All my friends given a good answer on Glucose estimation and I also have given my reply about two months back on the same topic. No doubt that DNS is a proven and routinely used procedure for glucose (reducing sugar) estimation. But this method has its own difficulties i.e.

1) DNS method is NOT SPECIFIC only for Glucose but for reducing sugars (may be -CHO group)

2) The time taken for estimations about 45 - 50 min. During the fermentation process one need to have `quick' result of left over `glucose' or sugar concentration, and if we get result after 45 - 50 min. one cannot feed the sugar/glucose to required level. Even if you feed the concentration would definitely affect the fermentation yield, growth, metabolic profile of the organism etc.

3) The only specific and quick method for the estimation of Glucose is by enzymatic estimation which gives quick result to run my process effectively.

Thanks

Dr.K.Prabakaran

I was learnt that DNS is NOT good for human health - it may be carcinogenic (needs to be confirmed)

Enzymatic kits are the best

Quikest and almost instantaneous method to detrermine glucose in fermentation samples is the one that is used for monitoring blood sugar (glucose) levels among diabetics using glucometers. This can be performed at regular intervals when sample is collected for determining optical density (OD) of culture, by taking a few micro-litres of the collected sample and adding it to the strip for determining the sugar levels (here patients blood sample is replaced by fermentation sample). Precaution should be taken to minimise the error by taking a constant volume in all the samples collected for this purpose and if necessary serial dilution may be performed, depending upon the concentration of glucose in a sample. This method can be applied to other reducing sugar compounds as the test is not very specific glucose. Methods like DNS and others are time consuming and cumbersome especially when one has to make adjustments in the feeding rate in order to sustain the exponential growth depending upon the amount of carbon source (glucose in this case) consumed. I have found glucometric method to be quick and efficient.

Dear Mr. Syed - the method which you have mentioned is very quick and specific to glucose but not to other reducing sugars and I have been using this for almost 15 years now; I have mentioned this method in my earlier mail about 2 months back. However, these strips are with glucose oxidase and hence specific for glucose depending on the source of enzyme.

The quicker method by using this analyzer, http://www.ysilifesciences.com/index.php?page=ysi-2700-select-analyzer

and for DNS refer to this paper: (A.Z.Sulaiman et al., 2011)

Sulaiman AZ, Ajit A, Yunus RM, Chisti Y. 2011. Ultrasound-assisted fermentation enhances bioethanol productivity. Biochemical Engineering Journal, Volume 54, page 141-150 (Impact factor = 2.645)

All the best....

DNS is the simplest known protocol

Enzymatic by HK-G6PDH / NAD to NADH2 / 340 nm

DNS _DiNitro Salycilic test is the right and fast method for glucose presence in fermentation broth.

DNS is the simple and rapid method...You may use GOD_POD kit for more accuracy

If the only reducing sugar you have is glucose I suggest the DNS method (Miller, 1959). Is quite reproducible and fast.

Cheers

I have done this in for both research and for my classes.

Miller, 1959 is the best reference for the DNS assay and it is easily scaled down to a 96-well plate method for very high throughput and you can easily link it to papers that discuss the methods various limitations and reasons behind the reagents components. there are a few papers that scale it to 96 well plates. It MUST BE USED WITH A STANDARD CURVE and if glucose/maltose (from starch hydrolysis) are preset ONLY, then it is accurate. other sugars have a minimal influence but notable.

GOD-POD and the glucometer strips are the best if glucose if is only present (you prepared the sugars, not sourced them from a hydrolysis) and if you can get the dilutions right the strips are fast. no fuss, no mess. Maltose will over estimate.

HPLC is only useful if you need to determine multiple compounds or you need to separate it from the matrix. HPLC is preferred if you have a complex sample and/or you want to determine ethanol at the same time.

Phenol-sulphuric is good for a pure sugar but it is most influenced by other sugars in the sample. Not to mention it involves high concentrations of a strong acid.

DNS or Somogy is the best and simplest method for determination of reducing sugars.

Check out http://www.cescobioproducts.com/gluccell-glucose-monitoring-system.html for a cheap and fast way to measure glucose in fermentation broth

GOD-POD method is the best

For accurate determination at a fixed time, it's first important to block fermentation in the sample.

Thanks for all of this, please wanted to ask something else, what can i use to stop the fermentation when drawing a sample...???

I successfully used the dilution of the sample with ethanol up to approx. 70% v / v , filtration prior to the determination by the enzymatic method.

Of course if you want to measure ethanol as well as glucose, as you always do in an ethanol fermentation, place the sample in a 1.5ml tube, centrifuge for a few seconds at top speed then filter the supernatant though a 0.45um filter, nylon preferred. shouldn't take anymore then a few seconds. The supernatant should be free of microbes for sugar determination. then you can also wash the cell pellet left over from the centrifuge tube and do dry mass or OD for biomass determination.

???

I used successfully a brix meter (use a pure glucose as standard to prior draw a regression slope)

for the second question I suggest Mashego et al; (2003) Critical evaluation of sampling techniques for residual glucose determination in carbon-limited chemostat culture of Saccharomyces cerevisiae he processed as follow: (i) quick freezing of fermentation broth directly in liquid nitrogen; (ii) quenching metabolism by exposing the fermentation broth to stainless steel beads (4-mm diameter) in a filter syringe precooled to −18°C; and (iii) withdrawal of the filtrate through a 0.45-μm filter attached to a syringe and a needle inserted directly into the fermentor.

Dear Mr. Roger - In my opinion the Brix meter may not give accurate result of glucose concentration in the fermentation broth, because the media components and microbes present in the broth would also interfere with the glucose reading. Hence, the Brix meter is usually not used in regular fermentation process and it is only used in Yeast fermentation when molasses is used

Dear Prabakaran

I completly agree with u about the acuracy of brix method that's why I precise "use a pure glucose as standard to prior draw a regression slope" as Mr Samani were looking for a"quicker method to quantify the amounts of glucose in a sample" I guest he's using pure glucose as substrate for bioethanol production.

The great discussion, also useful for me, I am also interesting that how to quantify H2O2 produced by glucose oxidation and I will do the GOD oxidation of glucose,

DNS can used for general reducing sugar analysis;

For exact glucose measurements, GOD POD assay is the right method.

A simple and relatively quick method to quantify glucose or any other reducing sugar is DNS.

DNS is the simplest way and quicker method plus cheaper cost

Fehling Method can be use. it will give you nearly correct % of glucose in media. it is faster n cheapper than other method.

GOD-POD method is the best method for glucose determination.

correction: Its GOD-PAP method, unlike DNS it do not need heating & just incubate only in room temperature, can order from Randox chemical

I meant GOD-POD method only, http://www.keegad.com/analytical-reagents/Gloucose_PackInserts.pdf

Mohd comment would be explicit if he could also give a link to GOP-PAP as Slvaraju has done for better comparism

You can use the DNS method if you have time but not funds, or an automatic enzymatic method (like YSI) if you have funds but not enough time between each fermentation point.

Recently, I am using hexokinase method as attached

I recommend using the enzymatic method for glucose determination with glucose oxidase

Simply use a glucose oxidase electrode.

Defend on sugar compose in broth fermentation. If intital substrate in broth fer. only glucose so DNS or Felling or somogy menthod are best suitable for fast determination.

Best regards

The fastest way is using biosensor with the membrane which can react with the glucose. You can read the result in 5 minutes.

Didi He, Is this biosensor commercially available?

You may visit www.cescobioproducts.com for their simple and affordable glucose meter which is designed for cell culture but has also been satisfactorily used for some microbial cultures if the medium is relatively clean or defined.

Why not to try Glucometer (Used for Diabetes), out of the box thinking !!

Cost efficient and fast...

Dns or god-pod....

Glucose (HK) assay kit (GAHK20-1KT) from Sigma maybe suitable for you.

use YSI 2900 Biochemistry Analyzer to determine amount of glucose, it is a fast and reliable method as compared to DNS method.

Glucose oxidase electrode....

Hello, use the glucose kit that are normally used in Medical labs.

You can use the glucose kit which give you the concentration of glucose present your sample within few minutes, also YSI glucose analyser can also be used. However, try to book the HPLC in advance so that you can analyse for other sugars as well.

Gluco test (GOD PAP) is easy and quick

DNSA, Gluco Test, KIts, glucose oxidase electrode,Somogy & HPLC etc. and several other methods are easily available.You want to analyze glucose contents to what accuracy?

as there are other cheaper and easy methods like Fehling are also available. You need to select a method with required accracy

During fermentation in shake I accompanied decay of glucose with glucose oxidase kit. It's fast. For other sugars HPLC - when there is time for it.

DNS method is the best solution to measure glucose concentration 

Colorimetric assays are much faster than HPLC and GC, particularly if you use them in a micro-plate format (96 samples simultaneously). Your options are the DNS assay or D-Glucose Assay Kit (GOPOD Format) - but please remember they assay different things. DNS measures the total reducing sugars, while GOPOD is specific to glucose.

Our lab recently optimized the DNS method precisely for this purpose (please see: http://www.sciencedirect.com/science/article/pii/S0961953412002097). The results in the paper allow you to tailor the DNS assay to get the most accurate quantification.

Hope it helps!

Diastix strips for fast but not so accurate measurement (1 min test better with the supernatant broth sample), blood glucose analyzer - small device like a computer mouse size with strips (enzyme reaction, 1 min test better with the supernatant sample) and it is very accurate up to 30 mM and can bought in a regular drug store

Greetings....

I am using GODPOD reagent for determining the D-Glucose. I am doing this determination with food stuffs. Food stuffs are giving high amount of glucose. For that, how to increase the linearity level of the GODPOD reagent to determine the high concentration of glucose without more dilutions of sample solution...???