scholarly journals A fixation procedure suitable for autoradiography of endogenous long-chain acyl carnitine.

1985 ◽  
Vol 33 (8) ◽  
pp. 744-748 ◽  
Author(s):  
M T Knabb ◽  
G G Ahumada ◽  
B E Sobel ◽  
J E Saffitz
Keyword(s):  
Subcellular Localization ◽  
Selective Extraction ◽  
Total Radioactivity ◽  
Water Soluble ◽  
Free Carnitine ◽  
Neonatal Rat ◽  
Short Chain ◽  
Tissue Processing ◽  
Chain Acyl ◽  
Long Chain

A tissue processing procedure was evaluated for fixation of endogenous long-chain acyl carnitine (LCA) to facilitate autoradiographic subcellular localization of this amphiphile. Suspensions of neonatal rat myocytes labeled with exogenous 14C-palmitoyl carnitine retained 85.2% of the radiolabel after tissue processing. Autoradiography demonstrated no significant translocation of radiolabeled LCA from myocytes to unlabeled sheep erythrocytes mixed in equal proportions and processed together. To evaluate endogenous LCA fixation, cultured myocytes were incubated for 3 days with 3H-carnitine. Radioactivity was distributed in LCA, short-chain acyl carnitine, and free carnitine pools in proportion to the physiological concentrations of the metabolites traced. Before tissue processing, LCA contained 4.5% of total radioactivity. After tissue processing, labeled water-soluble components were lost and 88% of the retained radioactivity was in the LCA pool. The enrichment of endogenous LCA radioactivity was attributable to the selective extraction of endogenous short-chain and free carnitine. Nearly 75% of endogenous LCA was preserved. In contrast, 99.5% of both endogenous short-chain and free carnitine were extracted. Thus, endogenous LCA can be selectively preserved, permitting quantitative subcellular localization of this amphiphile with ultrastructural autoradiography.

The lipase activity of a partially purified lipolytic enzyme of Escherichia coli

1978 ◽  
Vol 56 (5) ◽  
pp. 324-328 ◽  
Author(s):  
G. Nantel ◽  
Georgia Baraff ◽  
P. Proulx
Keyword(s):  
Escherichia Coli ◽  
Lipase Activity ◽  
Slow Rate ◽  
Water Soluble ◽  
Lipolytic Enzyme ◽  
Ph Optimum ◽  
Chain Acyl ◽  
Marked Preference ◽  
Hydrolysis Of ◽  
Long Chain

Escherichia coli lipase was found to have a broad pH optimum between pH 8 and 10. Long-chain acyl triacylglycerols such as trioleoylglycerol were hydrolysed at a relatively slow rate, whereas, the shorter-chain acyl derivative tricapryloylglycerol was not. Triacylglycerols and diacylglycerols were broken down at a rate 10- to 15-fold greater than that for monoacylglycerol. Simple esters such as methyloieate and cetylpalmitate were hydrolysed at rates greater than that of triacylglycerol. Water-soluble esters such as p-nitrophenylacetate were not attacked. Hydrolysis of lipase substrates occurred more readily in the presence of an anionic detergent such as taurocholate. The enzyme had no marked preference for the 1- or 3-position of triacylglycerols but attacked these positions much more readily than position 2. The enzyme also catalyzed transacylation reactions with simple alcohols such as methanol or ethanol.

Sources of error in determinations of carnitine and acylcarnitine in plasma.

1984 ◽  
Vol 30 (2) ◽  
pp. 316-318 ◽  
Author(s):  
R C Fishlock ◽  
L L Bieber ◽  
A M Snoswell
Keyword(s):  
Perchloric Acid ◽  
Volume Of Fluid ◽  
Fluid Volume ◽  
Free Carnitine ◽  
Short Chain ◽  
Distilled Water ◽  
Plasma Samples ◽  
Long Chain ◽  
Total Fluid

Abstract Radioactive and nonradioactive L-carnitine and acyl-L-carnitine were used to evaluate the washing procedures used during the determination of free, total, short-chain, and long-chain acylcarnitine in human and sheep plasma. The volume of fluid trapped by the protein precipitated by perchloric acid is approximately 24% of the total fluid volume and thus contains 24% of free carnitine and short-chain acylcarnitine. Washing twice with distilled water removes about 25% of the long-chain acylcarnitine along with the trapped free carnitine and short-chain acylcarnitines. Washing the pellet twice with a 60 g/L solution of perchloric acid completely removes the trapped free carnitine and short-chain acylcarnitine but does not remove the bound long-chain acylcarnitines. Thus washing with perchloric acid is essential for accurate measurement of long-chain acylcarnitines in plasma samples.

Acyl-CoA dehydrogenase activity in pea cotyledon tissue during germination and initial growth

2000 ◽  
Vol 28 (6) ◽  
pp. 760-762 ◽  
Author(s):  
C. Masterson ◽  
A. Blackburn ◽  
C. Wood
Keyword(s):  
Dehydrogenase Activity ◽  
Chain Length ◽  
Root Formation ◽  
Short Chain ◽  
Initial Growth ◽  
Chain Acyl ◽  
Pea Seeds ◽  
Acyl Coa Dehydrogenases ◽  
Hydroponically Grown ◽  
Long Chain

Acyl-CoA dehydrogenase activity has been measured in homogenates of post-imbibition to 14-day-old hydroponically grown pea seeds at daily intervals, using C4, C12 and C16 acyl-CoA substrates. The activity peaks of the different chain-length acyl-CoA dehydrogenases did not transpose at all points and the ratios of the chain-length activities were not constant. It therefore has to be concluded that more than one dehydrogenase is present in pea mitochondria. There was a post-imbibition initial surge of activity with short- and mid-chain-length substrates. The C16- handling enzyme first peaked at 3–4 days, which coincided with the onset of plumule unfurling and greening. Further peaks were observed with all three substrates, coinciding with secondary root formation and leaf enlargement and later with cotyledon degeneration. Overall activity showed that the long-chain acyl-CoA dehydrogenase was much more active than the short-chain acyl-CoA dehydrogenase.

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