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Metody oznaczania L-karnityny

Łącka A., Leszczyńska J.

Wiadomości Chemiczne

2002

[Z] 56, 3-4

341-360

L-carnitine (3-hydroxy-4-N-trimethylammonium butyric acid) plays an important role in fats metabolism, acting as an essential carrier of acyl group from cytoplasm into the mitochondria and b-oxidation products from peroxosomes. It is also responsible for detoxication of specific short- and medium-chain fatty acids. Although carnitine is not indispensable in food and can be synthesized in the system, its deficiency accompanies many diseases. Deficiency in carnitine is characterised by muscle weakness, cardiomyopathy, deposition of fat in muscles, kidneys, liver as well as metabolic acidosis. L-carnitine occurs in a free form and esterificated by fatty acids. For diagnostic purposes free and total carnitine are determined after previous ester hydrolysis. Modern methods of carnitine determining are based on reversible reaction of the stoichiometric acetyl-transfer from acetyl-coenzyme A to L-carnitine catalyzed by carnitine acetyltransferase (CAT), and measurement of the content of acetyl-carnitine or free coenzyme A formed. Isomer D-carnitine is inactive in that reaction, nor-carnitine (3-hydroxy-4-dimethylammonium butyric acid) may take part, but this compound does not appear in any biologic materials. Those methods of carnitine determining employ radiometric, spectrophotometric, spectrofluorimetric and enzymatic techniques. Among the other methods of carnitine determining chromatographic methods are most frequently used, especially HPLC, but also GC chromatography is often used. Using HPLC method, one can determine the carnitine level, either immediately or in the form of 4'-bromophenacyl derivatives. However, some authors have had difficulties in obtaining these derivatives and sensitivity of determination has been not sufficient for biological samples. Various reagents were used to prepare fluorescent carnitine derivatives in the HPLC chromatography with fluorimetric detection. The reactor containing immobilized enzymes was also used. One can also determine carnitine by means of gas chromatography. This method is based on the disintegration of carnitine at temp. 160 oC in the presence of NaOH and NaBH4. Under these conditions it is disintegrated into crotonylbetaine, trimethylamine and butyrolacton, which are detected with flame ionization. Among methods used less frequently mass spectroscopy and capillary electrophoresis are worth mentioning. Individual O-acyl derivatives content L-carnitine can be determined using the "radio-exchange" method or gas chromatography.