| The determination of serum triglycerides (TG) has long been an important clinical measurement in regard to identifying individuals with hyperlipidemia and at risk for cardiovascular diseases. Serum TG measurements are now performed in most clinical laboratories with enzymatic methods which are simple but susceptible to various sources of interference. Standardization of TG measurements needs reference methods. Current procedures for accurate measurements of triglycerides are chemical and isotope-dilution mass-spectrometric methods. The tedious sample preparation and expensive instrumentation involved in these methods prompted us to investigate the possibility of analyzing serum TG by high performance liquid chromatography (HPLC).Both methodologies with and without glycerol blanking have been used in enzymatic TG measurements. Thus, procedures intended for use as reference methods should be capable of analyzing at least two of the three analyte categories: total glycerol (defined as the sum of TG, di- and mono- glycerides and free glycerol), glycerides (TG plus di- and mono-glycerides or TG only) and free glycerol. Glycerol is the preferred molecule or molecule moiety that is to be analyzed in analyzing the above analyte categories. Glycerol is a saturated small molecule polyol and does not have adequate chromatographic separability and detectability on most HPLC systems. A pre-column derivatization is needed in HPLC analysis of giycerol. The pre-column derivatizations often used for alcohols are esterifications under anhydrous conditions with organic base as catalyst. But for glycerol in biological samples, both the evaporation of the extraction solvent and the purification of the derivative are difficult. It has long been known that hydrxyl compounds can also be benzoylated in aqueous... |
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