High-pressure liquid chromatography Sample preparation

Chocolate liquor is the solid or semiplastic food prepared by finely grinding the nib of the cacao bean. It is commonly called baking chocolate, unsweetened chocolate, or bitter chocolate and, in Europe, is frequently referred to as chocolate mass or cocoa paste. Chocolate liquor is essentially the starting point from which all chocolate products are produced. Table 5 lists the theobromine and caffeine content of 22 various chocolate liquor samples determined by high pressure liquid chromatography (HPLC). The liquors averaged 1.22% theobromine and 0.214% caffeine.27- 28 The ratio of theobromine to caffeine ranged from 2.5 1 to 23.0 1. 

Samples containing 150 g of 500 ppm acids were also prepared using 3% added sodium chloride in the 90/10 oil/water blend, water, and vegetable oil as the microwave medium. These samples were heated 0, 1, 2, and 3 minutes in the microwave. Changes in the acid concentration were determined by high pressure liquid chromatography with an organic acid column and an aqueous mobile phase. 

Analytical Techniques. Sorbic acid and potassium sorbate are assayed titrimetrically (51). The quantitative analysis of sorbic acid in food or beverages, which may require solvent extraction or steam distillation (52,53), employs various techniques. The two classical methods are both spectrophotometric (54—56). In the ultraviolet method, the prepared sample is acidified and the sorbic acid is measured at 250-260 nm. In the colorimetric method, the sorbic acid in the prepared sample is oxidized and then reacts with thiobarbituric acid the complex is measured at 530 nm. Chromatographic techniques are also used for the analysis of sorbic acid. High pressure liquid chromatography with ultraviolet detection is used to separate and quantify sorbic acid from other ultraviolet-absorbing species (57—59). Sorbic acid in food extracts is determined by gas chromatography with flame ionization detection (60—62). 

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. 

Osmotic effects play an important role mainly at high injected polymer concentrations. They may selectively affect the retention volumes of smaller polymer species contained in the polymer sample [97], Osmotic effects within porous column packings form the basis of interesting preparative liquid chromatographic method developed by Teraoka and coworkers [98-101] and denoted the high osmotic pressure liquid chromatography. 

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