Organic acids HPLC analysis

Chlorination of Individual Amino Acids. HPLC analysis of an extract of chlorinated humic acids indicated that the chlorination products compose a highly complex mixture of organic material. Thus, the task of identification of mutagenic products of chlorination would not be simplified by the use of the humic acid model. In contrast, the amino acid model of production of mutagenic compounds can be readily simplified by the use of individual compounds as precursors. 

In general, CE is simple, rapid, and low cost because it needs neither laborious treatment of the samples nor long times of analysis. However, its high detection limit is a major limitation of CE. CE is often poorly reproducible. Enzymatic assay is more suitable for quantifying one organic acid in honey samples because it is specific, precise, and accurate. GC is more suitable for analyzing volatile or semivolatile chemicals. HPLC is versatile and reproducible. However, common HPLC detectors such as UV-VIS are not very sensitive for organic aliphatic acids. 

For HPLC analysis of preservatives (antimicrobials such as polar organic acids, or antioxidants such as... 

With regard to AADC, vanillactic acid (a metabolite of 3-MD) appears in the urine and can be detected by organic acid analysis. As predicted, an increase in urinary vanillactic acid has also been reported in PNPO deficiency. The PLP concentration in CSF, which can be determined by HPLC, is also reported to be decreased in PNPO deficiency. Mutation analysis can also now confirm or refute a suggestion of PNPO deficiency. 

As stated in Basic Protocol 1, the method of HPLC used may change depending on the organic acids present in the sample. This method uses a Bio-Rad Aminex HPX-87H column for HPLC analysis. An internal standard is important for the analysis of organic acids. This provides a means of not only determining if the analysis is working but also quantitating the percent recovery of the method. 

Many methods have been used to determine organic acids in foods, including volumetric, electrochemical, enzymatic, and chromatographic (paper, thin-layer, gas-liquid, or HPLC) methods. However, most of these are not able to assay organic acids comprehensively for example, the enzymatic methods are specific kits for individual organic acids (i.e., they only detect one of the acids present). Therefore, to analyze the sample comprehensively using enzymatic methods would be extremely time consuming and costly, as the analysis would have to be run separately for each acid and would require several kits. 

The HPLC analysis for Basic Protocol 1 requires -50 min per run whereas Basic Protocol 2 requires -15 to 20 min per run. Once the standard solutions have been run for Basic Protocol 1 (-1 hr) or the standard curve has been made for Basic Protocol 2 (-2 hr), the sample can be analyzed. Sample analysis should be made in duplicate. After the HPLC analysis, the integration of the results and concentration calculations require -15 to 30 min per organic acid. 

KI Tomlins, DM Baker, IJ McDowell. HPLC method for the analysis of organic acids, sugars, and alcohol in extracts of fermenting cocoa beans. Chromatographia 29(11/12) 557-561, 1990. 

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