Fatty acids, radioactive

The fatty acids from each fraction were transesterified with methanolic-HCl (Stoffel et al., 1959), extracted by light petroleum, evaporated and assayed for fatty acid radioactivity in a Packard Scintillation Spectrometer. 

Table II Percentages of total fatty acid radioactivity in the fatly acids of different cellular fractions prepared from safflower cotyledons labelled with [1- C] linoleate.

Experiments with monkeys given intramuscular injections of a mineral oil emulsion with [l-14C] -hexa-decane tracer provide data illustrating that absorbed C-16 hydrocarbon (a major component of liquid petrolatum) is slowly metabolized to various classes of lipids (Bollinger 1970). Two days after injection, substantial portions of the radioactivity recovered in liver (30%), fat (42%), kidney (74%), spleen (81%), and ovary (90%) were unmetabolized -hexadecane. The remainder of the radioactivity was found as phospholipids, free fatty acids, triglycerides, and sterol esters. Essentially no radioactivity was found in the water-soluble or residue fractions. One or three months after injection, radioactivity still was detected only in the fat-soluble fractions of the various organs, but 80-98% of the detected radioactivity was found in non-hydrocarbon lipids. 

With highly purified enzyme, free from pyrophosphatase, radioactive exchange between 32PP and ATP could be demonstrated. Later it was shown that at least three different enzymes activated fatty acids, depending on the chain lengths of the substrate. 

Wolfe has presented an excellent description of the systematic application of stable and radioactive isotope tracers in determining the kinetics of intestinal fat absorption, hepatic triglyceride synthesis, lipid mobilization, triglyceride-fatty acid recycling, and cholesterol turnover. 

The inverse technique, which makes use of complexation of compounds to the product such that it becomes insoluble (e.g. fatty acids with barium cations), is also applied. These methods are also applicable for strains that do not grow on the screening compound due to lack of a complete metabolic pathway for the starting material. A variation on this theme is the trapping of radioactive tracers, such as... 

Pigs dosed with radiolabeled avilamycin produced three unidentified fecal metabolites derived from the oligosaccharide and/or the eurekanate moieties. However, the primary metabolite in feces and liver was flambic acid. Mean total residues in muscle were all below 0.2 ppm, whereas residues in other edible tissues were all below 1 ppm. Most of tissue residues were derived from the oligosaccharide and/or the eurekanate portion of avilamycin, whereas very little was parent avilamycin. In fat, the avilamycin related residues was found to be due to radioactivity that had entered normal metabolic pathways and had been incorporated into the fatty acids. 

Correct answer = D. Malonyl CoA (three carbons) is synthesized from acetyl CoA (two carbons) by the addition of CO2, using the enzyme acetyl CoA carboxylase. Because CO. is subsequently removed during fatty acid synthesis, the radioactive label will not appear at any position in newly synthesized fatty acids. 

A number of methods are available for following lipase activity. Although numerous modifications and variations have been introduced, the basic methods are (1) titration of the liberated fatty acids, (2) changes in surface tension, (3) colorimetric determination of the fatty acids, (4) use of gas-liquid chromatography, and (5) use of radioactive substrates. Kuzdzal-Savoie (1980) has reviewed the subject. 

In 1904, long before the advent of radioactive tracers, Knoop synthesized fatty acids labeled by chemical attachment of a benzene ring at the end opposite the carboxyl group. He prepared these compounds with both odd and even numbers of carbon atoms in straight chains and fed them to dogs. From the dogs urine he isolated hippuric acid and phenylaceturic acid, which are the amides of glycine with benzoic acid and phenylacetic acid, respectively. Knoop showed that the phenylacetic acid was produced from those fatty acids with an... 

Normal human fibroblasts grown in media with 12% fetal calf serum were treated with or without galactose oxidase and then subjected to reduction with potassium borotritiide. GSLg were isolated and subjected to acid-catalyzed methanolysis (62) (see Materials and Methods). Methyl glycosides, methyl sphingosine, and fatty acid methyl esters were isolated by extraction of the total hydrolysate with solvents as described previously (62). Radioactivity was measured in triplicate in aliquots of these extracts. 

We were also able to use FAB mass spectrometry to determine the amino acid sequence around the active site serine in the acyl transference domain of rabbit mammary fatty acid synthase.6 The synthase was labelled in the acyl transferase domain(s) by the formation of O-ester intermediates after incubation with [" " C]-acetyl- or malonyl-CoA (Fig. 2A). The modified protein was then digested with elastase (Fig. 2B), and radioactive material isolated via successive purification steps on Sephadex G-50 and reverse phase HPLC. The isolated peptides were then sequenced by FAB MS. The data summarized in Table II established the sequences of both the acetyl and malonyl hexapeptides to be N-acyl-Ser-leu-Gly-Glu-Val-Ala. 

On the other hand, some fish are able to synthesize long-chain polyenoic fatty acids (Kayama et al., 1963) from shorter carbon chains. Docosohexaenoic acid is laid down in coho salmon in quantities related to the size of the fish, rather than to its availability in the diet (Tinsley et al., 1973). Rainbow trout fed on 18 2 and 18 3 fatty acids can produce 20 3, 22 5 and 22 6 fatty acids in substantial quantities (Owen et al., 1975), but these workers noticed that the capacity of marine flatfish to elongate or desaturate the carbon chains was more limited. They found that 70% of the radioactivity of labelled 18 3 appeared later in the 22 6 fatty acid of rainbow trout, but that turbot converted only 3-15% of labelled precursors into polyunsaturated fatty acids of longer chain length. It was suggested that turbot in the wild probably received adequate polyunsaturated acids in their diet, which the fish therefore did not need to modify. The elongation of the carbon chains and the creation of more double bonds is also only slight in Atlantic cod, another marine teleost, presumably for the same reason (Ross, 1977). 

In the past, the fish used as food for man or farm animals was evaluated on the basis of protein and total lipid content, but nowadays the emphasis is on the biologically active substances such as polyenoic fatty acids, vitamins and antioxidants. Recent problems have arisen because of contamination with sewage, mineral oil and its by-products, heavy metals and radioactive pollutants. The bizarre finding of certain Tasmanian oysters, bred downstream from a zinc smelting plant, which contained 10% of zinc on a dry weight basis is an... 

Chemiluminescence is a very sensitive and selective technique. Reagent types, analytes, and detection limits have been summarized in a review by Imai.56 Chemiluminescence has been applied to the analysis of compounds that exhibit low UV absorbance, including metal ions, amino acids, fatty acids, and bile acids. Other detectors include detectors for radioactivity, nuclear magnetic resonance (NMR), and surface-enhanced Raman spectroscopy. Radioactivity detection is one of the most selective detectors, as only components that have been radiolabeled will be detected. The interface of NMR with HPLC and has been discussed in detail by Grenier-Loustalot et al.57 Surface-enhanced Raman spectroscopy is another technique that... 

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