The characterisation of docosahexaenoic acid (22:6n-3) biosynthesis in the Liver of rainbow trout (Oncorhynchus mykiss). - PhDData

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The characterisation of docosahexaenoic acid (22:6n-3) biosynthesis in the Liver of rainbow trout (Oncorhynchus mykiss).

The thesis was published by Buzzi, Marcelo, in September 2022, University of Stirling.


The pathway for the biosynthesis of docosahexaenoic acid (22:6n-3) fromlinolenic acid (18:3n-3) was investigated in rainbow trout (Oncorhynchus my kiss)liver in vitro, using primary cultures of hepatocytes and liver microsomes toinvestigate the products of desaturation and elongation of [l-14C]-18:3n-3 and[l-14C]-20:5n-3. Argentation thin-layer chromatography and radio gaschromatography were employed to analyse the metabolic products of the radiolabelledfatty acid substrates and to determine the rate of conversion of 18:3n-3 to 22:6n-3. The recovery of radioactivity in various polyunsaturated fatty acids (PUFA) of trouthepatocyte lipid, including 20:5n-3 and 22:6n-3, established that juvenile trout werecapable of converting 18:3n-3 to 22:6n-3. To establish the extent to which theformation of 22:6n-3 was enhanced in the absence of dietary PUFA, particularly20:5n-3 and 22:6n-3, and therefore, facilitate the investigation of 22:6n-3biosynthesis, rainbow trout were fed a diet based on olive oil and deficient in (n-3) PUFA. Feeding the diet deficient in (n-3) PUFA efficiently reduced the endogenouslevels of (n-3) PUFA in trout hepatocytes and at the same time markedly increased therate of 22:6n-3 formation from both [l-14C]-18:3n-3 and [l-14C]-20:5n-3. When thedesaturation and elongation of [1-14C]-18:3n-3 and [l-14C]-20:5n-3 were investigatedin microsomes isolated from trout liver, no radioactivity from either substrate wasrecovered in 22:6n-3. High proportions of radioactivity from [l-14C]-20:5n-3 were,however, recovered in 24:6n-3 and 24:5n-3. These radiolabelled C24-PUFA producedby the microsomal incubations were separated by argentation chromatography andused as substrates in incubations with hepatocytes isolated from trout liver. Docosahexaenoic acid (22:6n-3) was generated from both radiolabelled C24-PUFAsubstrates by trout hepatocytes. The results establish that the biosynthesis of 22:6n-3in rainbow trout hepatocytes does not involve A4-desaturation of 22:5n-3 but ratherproceeds via the A6-desaturation of 24:5n-3 with the subsequent chain shortening ofthe 24:6n-3 produced. Cyclopropene fatty acids and curcumin significantly inhibited the desaturation and elongation activity of hepatocytes and liver microsomes fromrainbow trout. The A6 and A5-desaturase and elongase substrate specificities wereinvestigated; it was shown that (n-3) PUFA substrates were always preferred by theenzymes over (n-6) PUFA.

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