Well, it is not my professional field. Unfortunately, I have no idea about it. Sorry for that

It is possible by taking the mass spectrum of the compound and comparing the fragmentation pattern with that of authentic . It may be there in Merk Index. You may also be able to find the lamda max in UV in Merk Index.

All the best

Anil

Determining the concentration of free ABA is also not very difficult

Thank you anil, do you have a published method as reference??

Finding a UV-Vis trace for ABA-GE is not going to be easy, as the techniques have moved on now and mass-spec is likely to be the most popular analytical method in the literature nowadays (apart from immunoassays). An alternative for you (assuming you can quantify losses) would be to hydrolyse your purified extract to release free ABA and then to assay total free ABA before and after hydrolysis.

The early literature on ABA-GE dates mainly from the 70s and 80s of last century, and you may find a UV-Vis spectrum for ABA-GE amongst these early papers - try a search for "abscisic acid" + glucoside or glucosyl ester and Milborrow or Abrams or Zeevaart. These three have been key players in the ABA field and will have used physico-chemical methods to characterise ABA and its metabolites.

There could be a UV spectrum of ABA-GE in one of the early books on ABA, such as "Abscisic acid" (ed F. T. Addicott), Praeger, New York (1983).

This begs the question of why you want to measure ABA-GE just because there is a lot more of it in Vitis vinifera leaves than free ABA. Unless ABA-GE has some sort of functional role in Vitis vinifera, then the question could be asked: why do you need to measure it? Someone who did extensive work on ABA in grape was BR Loveys in the 1980s, and later.

ABA glucose ester certainly used to be available from chemical suppliers, though I suspect the demand for it now doesn't justify its manufacture.

You can obtain an ABA-GE standard from OlChemIm ltd (Olomouc, Czech Republic) for an affordable price.

Dear Alessandra,

Greetings!

I think the lambda max for ABA-GE is 265 nm. I am herewith enclosing an article for your kind perusal. I hope it will be of some help to you.

Take care

With Regards,

Rajeev K Singla

SERB-Young Scientist

[email protected]

http://rajeevksingla.in

The reference from Rajeev K Singla does not give any UV information on ABA-GE, but it does give references to earlier work where you will find the use of UV spectroscopy to record ABA-GE. One of the references is by Neill, Horgan and Heald, Plant 157: 371-175 (1983): http://link.springer.com/article/10.1007/BF00397410#page-2.

Here you will see that UV maxima for ABA-GE were located at 273 and 240. The peak at 273 is probably much larger than the one at 240. To quantify this, though, you will also need the molar extinction coefficient, which may or may not be given later in the Neill et al. reference (not available to me).

If you can access it, the original paper describing ABA-GE is KOSHiMIZU K, M INUI, H FUKUI, T MITSUI, 1968. Isolation of (±)-abscisyl-,l-Dglucopyranoside

from immature fruit of Lupinus luteus. Agric Biol Chem 32:

789-791.

I hope that's some help.

Cheers,

Steve

Thank you; I read the reference proposed by Rajeev (Boyer, 1982) so I knew there were no indications about the UV-VIS spectrum.

Your indications are precious, I will look for the manuscripts you have indicated but as they are old I am not sure to be able to obtain them.

My problem as to chromatograms is the following: besides isolating the free ABA (that I have identified with retention time and correspondance with the spectrum of the pure molecule) I isolate other molecules which could be ABA-GE; their spectrum of absorbances has two peaks: one (the higher) around 270 nm and the other around 345/350 nm. Could this second peak be attributable to glucose of the glycosid form?

I am afraid not. With absorbance maxima at 270 and 345/350 nm, I would suspect a flavonoid or related compound.

ABA-GE could be synthesized by the help of any organic synthesis of your colleagues.

You might also try Evaporating Light Scattering Detector (ELSD), it is very powerful with poor UV absorbing molecules. It could be used in separate or with HPLC.

Regards

The spectra of abscisic acid and the glucose ester are similar at pH less than 4 with the peak of the GE red shifted to 273nm. They both have a peak at 265 and 273 respectively and shoulder at 240. Above pH 7 the maxima of abscisic acid shifts from 265 to around 250, while the glucose ester remains the same as in acid pH. Boyer and Zeevaart 1982 Plant Physiol give an extinction coefficient of 20000 at 273nm for the GE in methanol. There is no peak at 350nm.

Thank you a lot for your really helpful information

dear Gaetan,

me too I suspected a flavonoid (flavonol in particular with absorbance at about 360 nm). I injected a standard solution of free Aba dissoved in methanol and I obtained the expected spectrum; threafter I submitted an aliquot of free ABA standard solution to the same steps of extraction as samples and I obtained two peaks in the chromatogram with two maxima absorbances, one around 260 nm and the other around 350 nm. So I think that these peaks could be reaction products of free ABA submitted to the numerous and tricky steps of ABA extraction. Could this be likely?

Thank you lot

Alessandra

Dear Alessandra,

Depending on your extraction conditions, it is possible that ABA degrades, but it is impossible that ABA-GE is formed from free ABA during mock extraction, unless you added a glucosyltransferase to the tube! The second peak could also be a contaminant retrieved during extraction. Do you still have one or both peaks with a blank sample submitted to extraction? For more information on the nature of that second peak, you may use mass spectrometry if you have access to it. ABA-GE gives three main ions at m/z 425, 263 and 153 in negative mode.

dear Gaetan,

I did not add glucosyltransferase to the tube.... and I have no peaks in the blank...

I have 2 ways now: don't care about these peaks (at the end my goal is measuring free ABA) or find an acces to a LC-MS; thank you for indicating the m/z of the main ions.

grazie

alessandra