Amines, determination extraction clean

Spengler and Jumar [90] used a spectrophotometric method and thin layer chromatography to determine carbamate and urea herbicide residues in sediments. The sample is extracted with acetone, the extract is evaporated in vacuo at 40°C and the residue is hydrolysed with sulphuric acid. The solution is made alkaline with 15% aqueous sodium hydroxide and the liberated aniline (or substituted aniline) is steam distilled and collected in hydrochloric acid. The amine is diazotized and coupled with thymol, the solution is cleaned up on a column of MN 2100 cellulose power and the azo-dye is determined spectrophotometrically at 440nm (465nm for the dye derived from 3-chloro- or 3.4-dichloroaniline) with correction for the extinction of a reagent blank. 

For precise quantification of lower dosed drugs in plasma, another workup was necessary to improve the signal-to-noise ratio (S/N). Since only a limited number of quite similar compounds had to be isolated, SPE was preferred, resulting in rather clean extracts. However, to compensate for the known batch-to-batch differences of SPE columns [29], deuterated analytes were used as internal standards. Heptafluorobutyric anhydride derivatization was preferred for primary and secondary amines to reach the sensitivity necessary for determination of low blood levels. 

Amines and amino acids were extracted as described by Tachiki and Aprison (1975). Tissues were homogenized in 2 ml of acidified-n-butanol and centrifuged at 1500 x g for 30 min. (4 C). A 1.0 ml aliquot of supernatant was returned to a second set of tubes containing 4.0 ml n-heptane and 0.4 ml O.IN HCl. This mixture was shaken for 10 min. and the layers cleanly separated by centrifugation at 1000 x g for 5 min. The organic layer was removed and 5-HIAA content was determined by the OPT fluorometric method. Serotonin (in the O.IN HCL) was assayed by the OPT fluorometric method while tryptophan was determined fluorometrically as described by Denckla and Dewey (1967). Tyrosine was determined flurometrically by the method of Waalkes and Undenfriend (1957). 

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