Organic acids derivatization techniques

Offline precolumn derivatization is the most common alternative in this respect it involves separating the esters obtained from the organic acids by reversed-phase chromatography, which amply surpasses solvophobic chromatography (i.e., the use of undissociated acids as such) and allows gradient elution techniques to be applied, thanks to the wider lipophilicity range covered by the derivatized compounds. 

Liquid absorption is a common technique for enriching compounds in reactive liquids like solutions of dinitrophenylhydrazine (DNPH) (for aldehydes), acetyl acetone (for formaldehyde) or aqueous carbonate solutions (for organic acids), both procedures which combine trapping and derivatization of the target compound. Another possibility is the use of dissolved alkali or acids to trap certain substances by the formation of salts in the solution. 

Ion chromatography is a special technique which was developed for the separation of inorganic ions and organic acids. The common detection principle is the monitoring of eluate conductivity although some analytes can also be detected with UV or, perhaps after derivatization, with visible light. Typical applications are the analysis of ... 

Clark, T.J. and J.E. Bunch Derivatization/headspace techniques for the analysis of organic acids in tobacco ... 

Basic techniques for speciation analysis were developed in the early 1980s they generally involve various analytical steps, e.g. extraction either with organic solvents (e.g. toluene, dichloromethane) or different acids (e.g. acetic or hydrochloric acid), derivatization procedures (e.g. hydride generation, Grignard reactions), separation (GC or HPLC), and detection by a wide variety of methods, e.g. atomic absorption spectrometry (AAS), mass spectrometry (MS), flame photometric detection (FPD), electron capture detection (BCD), etc. Each of these steps includes specific sources of error which have to be evaluated. 

Application of gas chromatographic techniques is restricted by the necessary volatility that the analyzed compounds should exhibit. A derivatization reagent is added, in a prior step, to render volatile the components of the analyzed organic materials. In some cases, a prior step consisting of an acid or alkaline hydrolysis of the organic material is necessary for releasing the molecular constituents of the polymeric structure. Inclusion of a prior step in the preparation procedure devoted to the suppression of interfering species is sometimes included. 

Isolation of the feeding factor for M. sexta was a far more difficult task. Whereas 2-tridecanone is a simple, stable molecule soluble in organic solvents, the feeding factor is water soluble, occurs at trace levels in plant tissue, and is easily hydrolyzed under mild alkaline or acidic conditions with subsequent loss of biological activity. The isolation of such a compound was a formidable obstacle requiring a departure from the more classical approach of hydrolysis or chemical derivatization followed by isolation of the lipophilic product. The necessity that pure substance be isolated with retention of biological activity required some basic research in modem separation techniques to develop a suitably mild isolation strategy. 

The most common analytical technique for the analysis of FFAs and their breakdown products has been chromatography. HPLC has been used for the analysis of FFAs (Christie, 1997 Lues et ah, 1998 Zeppa et ah, 2001). Analysis of short-chain fatty acids (C2-C4) may be relatively simple (Zeppa et ah, 2001). However, the analysis of long-chain fatty acids (>C6) may require derivatization. They are extracted using solvents, converted to bromophenacyl esters, and analyzed by reverse-phase HPLC. GC (with sample preparation and derivatization) has been the method of choice for analysis of fatty acids. An ideal but difficult procedure is to extract FFAs from the aqueous phase and organic phase and combine them (IDF, 1991). The challenge is to overcome the effects of partitioning and extraction of compounds that interfere with the analysis. ISO and IDF have detailed some of the extraction methods for lipids and liposoluble compounds in milk products (ISO, 2001b). Several other methods, which are mainly different in the extraction and derivatization steps, were reviewed by Collins et ah (2004). 

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