Liquid chromatography HPLC comparison

Lanina et al. 1992 Oishi 1990). These methods include gas chromatography (GC) combined with mass spectrometry (MS) and high-performance liquid chromatography (HPLC) combined with an ultraviolet detector (UV). No comparisons can be made between methods since no data were given regarding sensitivity, recovery, or precision. 

In high-performance liquid chromatography (HPLC) pressure is applied to the colunm that causes the solution to rapidly pass through it, allowing procedures to be completed in a fraction of the time in comparison to regular chromatography. 

Dimethylamine (109), putrescine (111), and spermidine (110), isolated from various insects (Table VIII), were obtained as p,p -nitrophenylazobenzoyl, p-phenylazobenzenesulfonyl, and I-dimethylaminonaphthalene-5-sulfonyl (dan-syl) derivatives and picrates or were detected by high-performance liquid chromatography (HPLC) using the ion-exchange resin (106,343). V,V-Dimethyl-3-phenylethylamine (131) from spiders of the genus Sclerobunus (Table VIII) has been identified by mass spectral comparison with a synthetic sample (117). 

Fig. 4.3. High performance liquid chromatography (HPLC) of the monosaccharides obtained from a partially purified preparation of microbubble glycopeptide surfactant from forest soil. Following hydrolysis (in 2 N HC1 for 6 hr at 100°C) and filtration, the carbohydrate mixture was charged on a Bio-Rad HPX-87 cation exchange column. For comparison, part A shows the chromatogram (using the same HPLC column) of a standard solution, which contained 4 pg of each of three different monosaccharides (i.e., the last three peaks shown are glucose, xylose and fiicose, in the order of increasing retention times). Part B shows the chromatogram obtained from hydrolysis of the partially purified (see text) microbubble surfactant (approximately 30 pg). All other experimental conditions were identical in the two cases, i.e., water eluent, 0.5 ml/min flow rate, 85°C, refractive index detector attenuation -2x. (Taken from ref. 322.)...

The GC is often connected to a mass spectrometer. Mass spectrometry (MS) breaks samples apart and separates the ionized fragments by mass and charge. Vast libraries of comparison fragments make computer-aided identification of materials possible even when the sample is very small. Most forensic laboratories have access to a combined gas chromatograph/mass spectrometer (GC/MS). High pressure liquid chromatography (HPLC) separates many types of drugs and may also be combined with MS. 

A comparison of the results obtained by high pressure liquid chromatography (HPLC) with results obtained by other techniques such as inductively coupled plasma emission spectroscopy (ICPES) showed good agreement and HPLC procedures are found to be economical in terms of analysis time. 

Chromatographic methods (1-25), in particular thin-layer chromatography (TLC) (5-17, 20-25) and more recently high-performance liquid chromatography (HPLC) (21, 22, 25), preferably are used. If dye mixtures are present, as is the case with madder dyeings, chromatographic comparisons are the best method of distinguishing the individual components. 

Preferably, high pressure liquid chromatography (hplc) is used to separate the active pre- and cis-isomers of vitamin D3 from other isomers and allows theic analysis by comparison with the chromatograph of a sample of pure reference j-vitamin D3, which is equilibrated to a mixture of pre- and cis-isomers (82,84,85). This method is more sensitive and provides information on isomer distribution as well as the active pre- and cis-isomer content of a vitamin D sample. It is applicable to most forms of vitamin D, including the more dilute formulations, ie, multivitamin preparations containing at least 1 lU/g (AOAC Methods 979.24 980.26 981.17 982.29 985.27) (82). The practical problem of isolation of the vitamin material from interfering and extraneous components is the limiting factor in the assay of low level formulations. 

A stable isotope dilution assay using mass spectrometry to measure insulin, proinsulin, and C-peptide has been developed. The difference in mass among the three analytes allows specific measurement of each protein. Comparison of patient samples revealed that most, but not all, results were higher by immunoassay than mass spectrometry. Thus immunoassays may overestimate insulin, particularly at low concentrations. The high protein concentration in the serum requires extraction of proteins (e.g., by immunoaffinity) and purification by high-performance liquid chromatography (HPLC) before quantification by mass spectrometry. This method is not suitable for routine laboratory analysis. 

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