ANALYSIS OF GLUCOSE BY HPLC

[afiqahtarmidi]

i need to know besides using hplc-rid, can we used other types of detector to detect glucose content in palm oil trunk?

help me...help me...

[Consumer Products Guy]

Evaporative light scattering detector (ELSD).

Sounds like you have UV-vis as your only HPLC detector.

1. Do you also have GC available ?

2. Can you use a UV-derivatization agent ?

[oscarBAL]

Glucose in palm OIL???
I supose you need sencitivity right?
Do you have the HPLC already and now are you loonking for the detector?
for some aplication people use ECD, elechtroquemical detector also known as voltammetric detector.

[HPLCCONSULT]

As I recall, sugar concentrations in some types of palm oil, Coconut Palms for example, is rather high (~ 7%) so detection may not be a problem. A basic search on the Internet will turn up plenty of papers on the topic. Many types of detectors should work well here.

Your question can be easily answered if you define what type of palm oil you are interested and then determine what concentration range is present (the data can be found on the Internet). That info will allow you to make an intelligent choice regarding which types of detectors and methods will work.

[JGK]

We found this quite useful http://www.waters.com/webassets/cms/lib ... A64081.pdf

We don't have UPLC but adapted accordingly

[oscarBAL]

HPLCCONSULT; thanks you very much for your explanation!!! now I am a little less ignorat in terms of palm oil
I was looking in the internet and I found a lot of information; metrohm have nice applications for sugar analysis using ECD or PAD (Pulsed amperometric detector not voltammetric as I stated before).

Yes, palm oils can have quite high level of sugar; afiqahtarmidi, sorry for ask but I would like to know why you need other detector beisde RID? do you only only need to analyze glucose or other parameter? I am just curious.

[unmgvar]

the standard reason for the develoment of other types of detectors for sugars and every other of non-UV detectable molecules is for several reasons:
1. RI is a universal detector that see all, but with very bad sensitivity. RI cannot be used with gradients so also it is more difficult to get chromatographic separation only in isocratic modes, that also for long runs with diffusion of peaks again bring back the problem of sensitivity
2. you could use fluorescence detection, which allows for gradients and give a very good sensitivity, but also, another step for sample preparation, more errors, signal also can change over time for the samples, and in some cases the fluo add-on actully effects the separation and causes the method to be hard to get.
3. MS can be used also but does everybody has the budget for this?

i recently dealt with a sugar application for wines where there is no problem of sensitivity for the sugars in red wine but simply other stuff as well that needed to be separated and with only isocratic mode it can be challenging

[afiqahtarmidi]

thanks for all the responders..

i tried too many times to search for others detector besides RID..
im already tried ELSD but then the peak is not suitable..
i just want the simple instrumentation as this is just my minor research.. for 2-3 days before i found we can used uv-vis spectrometer to detect the amount of sugar with some calculation.. but then hv to add bromothymol blue..
anybody here, can i used another types of this indicator?

really appreciated for your respond. at least i have some views about this research

[afiqahtarmidi]

dear oscarBAL:

my rid is not stable n i dun know what the prob it is.. becoz that i want to change to another detector as simple as that.. besides glucose, i try to detect the xylose also.. only these two types of sugar..
it is quite difficult to me because i dun have a guide very much..i have to find abt this my self. anybody can help me is welcome..
im to confusing abt this.

[tom jupille]

Before anyone can give you specific advice, you have to share details about what you are doing.
What kind of column?
What mobile phase (solvent system)?
What temperature?
What problem exactly (baseline drift? Noisy baseline? No peaks at all?)
Is the problem consistent or intermittent?

[afiqahtarmidi]

tom:

this is the ans for uy questions

What kind of column?
this column is Revex RPM-Monosaccharide Pb2+ (8%). all the samples must be in neutral (pH=7) b4 we run the experiment

What mobile phase (solvent system)?
for this column, i just can used distilled water becoz this is stated in broacher given to us

What temperature?
40 0c

What problem exactly (baseline drift? Noisy baseline? No peaks at all?)
i have the prob abt noisy, baseline n all that.the peaks r appeared but then the retention time of standard is keep changing even though we used the same standards. the prob also same when we tried to prepare a new standards.
the peak also broad..may hve overlapping of peak. i already tried to do the dilution but then still the same.
so, what can u suggest to me to improve this prob?

Is the problem consistent or intermittent?
the prob consistent day by day..

[lmh]

you can do sugar by UV-vis spectrophotometry with considerably higher specificity than you get from indicators by doing an enzyme coupled assay, but it's a bit more work than the indicator method, and still involves some calculations! And it's harder to extend it beyond the obvious sugars that are substrates for enzymes easily obtained from chemical companies (i.e. glucose, fructose, sucrose etc.). If you've got the equipment, and only need these, I still think it's less work than chromatography.

[tom jupille]

Much as I hate to admit it, lmh has a point: HPLC is not necessarily the best approach to all problems.

That said, the fact that your retention time is changing tells me that the detector is not the primary cause of your problems (the detector has no effect on retention time). Given a straightforward mobile phase like water, the two prime suspects are flow rate and temperature. RI detectors are sensitive to both.

Try monitoring the system pressure to see if it varies synchronously with your baseline. If it does, that confirms a flow problem. Make sure you are degassing properly and clean and/or change the check valves. You should also contact the pump vendor and ask them to provide a performance qualification (PQ) procedure so that you can verify that the pump is in fact operating properly.

Temperature is the other possibility. If you can, clip a thermocouple to the column and monitor the actual temperature (even better would be to put a thermocouple in a "tee" fitting immediately before the detector) and check the temperature (as with flow, what you are looking for is changes that sync with your basline variations). If your system is set up such that the transfer line from the column outlet to the detector inlet runs open to the atmosphere, make sure that line is insulated (a poorly insulate line can contribute to RI detector noise, but should not affect retention).

[venuzz]

Maybe you can try to increase your temperature and check for the stability of your temperature.
I ever use the same characteristic column as yours, I used Varian Hiplex Pb and it need to increase the temperature to about 80 celcius degree to separate sucrose, lactose, glucose and fructose. I used this column to separate the sugar content in coffee and coffee mixture.