Dear Dr. Olorunfemi Eseyin,

The analysis of triterpenoid/steroid saponins by GC-MS is not a suitable technique in my opinion. The main issue is the volatility of the compounds. The sapogenins obtained by acid or enzymatic hydrolysis is still not volatile enough to identify them by GC-MS aided by library search. While the individual sugar units can be identified from their trimethylsilyl derivatives. If more than one type of sugar units are there, their sequencing is the second issue. The third issue is the position and stereochemistry

of the sugar units in the aglycon. But otherwise if you have 15-20 mg of the pure saponin, the structure can be identified by 1D and 2D NMR and HRFABMS. Plenty literature on C NMR data is available . It is possible to get computed aided literature search to make life easier.

Only thing to worry about to avoid the artifact formation. You can get best separation by using reverse phase chromatography on modified silica gel such asODS or OS.

Dear Dr. Olorunfemi,

I had been working with triterpenes for about 4 years and had been using GC/MS to analyze them. In my experience, GC/MS just does not work well with intact triterpenoids. To this aim, I think that LC/MS would probably work better.

As you pointed out, hydrolysis can be carried out either under mild and strong conditions. There is this paper by J. Gutterman you might be familiar with.

This might help.

"Isolation and Structures of Avicins D and G: In Vitro Tumor-Inhibitory

Saponins Derived from Acacia victoriae"

Gamini S. Jayatilake,*,† Delano R. Freeberg,† Zhengjie Liu,† Steve L. Richheimer,† Mary E. Blake (Nieto),‡

David T. Bailey,† Valsala Haridas,§ and Jordan U. Gutterman§

J. Nat. Prod. 2003, 66, 779-783

I agree with both of the above responses and would say that water would be a poor solvent for injection due to its large expansion volume, personally I would do it in methanol if I was going to try it (I don't have much of a reason for preferring methanol to dioxane.)

The volatility is your primary concern, please provide more details about the matrix of your sample. I think getting an NMR of your sample would be a great first step, since you need such a small sample amount and because it will compliment the MS data should you choose to try analyzing with GCMS.

Would LCMS be available to you? What methods have you considered for purifying the different saponins if you do have several in the matrix (centrifuge, LC w/ fraction collector)?

Dear Dr. Olorunfemi Eseyin,

Saponins are highly polar compounds and non- volatile in nature. Apart from GC-MS, LC-MS is better for analysis for saponins.

LC-MS is better for analysis for saponins.