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1.
[(14)C]Streptozotocin was synthesized specifically labelled at three positions in the molecule. The biological activity of synthetic streptozotocin was characterised by studies in vivo of its diabetogenic activity and its dose-response curves. After this characterization the excretion pattern of all three labelled forms of streptozotocin was studied. With [1-(14)C]streptozotocin and [2'-(14)C]streptozotocin the injected radioactivity was excreted (approx. 70% and 80% respectively) mainly in the urine, the greater part of the excretion occurring in the first 6h period; small amounts (approx. 9% and 8% respectively) were found in the faeces. In contrast, with [3'-methyl-(14)C]streptozotocin a much smaller proportion (approx. 42%) of the injected radioactivity was excreted in the urine, the major proportion appearing in the first 6h, whereas approx. 53% of the injected radioactivity was retained in the carcasses. In whole-body radioautographic studies very rapid renal clearance and hepatic accumulation of the injected radioactivity was observed with all three labelled forms of the drug. There was some evidence for biliary and intestinal excretion. Major differences were apparent in the tissue-distribution studies, with each of the three labelled forms, particularly with [3'-methyl-(14)C]streptozotocin. There was no accumulation of [1-(14)C]streptozotocin in the pancreas for the 6h period after administration. However, with [3'-methyl-(14)C]streptozotocin (and also [2'-(14)C]streptozotocin) there was evidence of some pancreatic accumulation after 2h. The results indicate that streptozotocin is subjected to considerable metabolic transformation and to rapid renal clearance. The implication of these suggestions is evaluated with particular reference to the diabetogenic action of streptozotocin. 相似文献
2.
Differences in adult and foetal human cytochrome P-450 forms recognized by monoclonal antibodies with specificity for the P450III family.
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The biosynthesis of 9-[5'-deoxy-5'-(methylthio)-beta-D-xylofuranosyl]adenine (xylosyl-MTA), a naturally occurring analogue of 5'-deoxy-5'-methylthioadenosine (MTA) recently characterized, was studied in the nudibranch mollusc Doris verrucosa. Experiments performed in vivo with putative labelled precursors such as [8-14C]adenine, [Me-14C]methionine and [Me-14C]MTA indicate that xylosyl-MTA originates from MTA. Experiments with MTA double-labelled at critical positions are consistent with a 3'-isomerization of the nucleoside through the formation of a 3'-oxo intermediate. In addition, experiments with the newly synthesized [3'-3H]xylosyl-MTA are indicative for a very low turnover rate of this molecule, which therefore accumulates in the mollusc. 相似文献
3.
Metabolism of sialic acids from exogenously administered sialyllactose and mucin in mouse and rat 总被引:1,自引:0,他引:1
A mixture of N-acetyl-[4,5,6,7,8,9-14C]neuraminosyl-alpha (2-3(6]-galactosyl-beta (1-4-glucose[( 14C]sialyl-lactose) and N-acetylneuraminosyl-alpha (2-3(6]-galactosyl-beta(1-4)-glucit-1-[3H]ol(sialyl-[3H]lactitol) as well as porcine submandibular gland mucin labeled with N-acetyl- and N-glycoloyl-[9-(3)H]neuraminic acid were administered orally to mice. The distribution of the different isotopes was followed in blood, tissues and excretion products of the animals. One half of the [14C]sialyl-lactose/sialyl-[3H]lactitol mixture given orally was excreted unchanged in the urine. The other half was hydrolysed by sialidase and partly metabolized further, followed by the excretion of 30% of the 14C-radioactivity as free N-acetyl-[4,5,6,7,8,9-14C]neuraminic acid and 60% of this radioactivity in the form of non-anionic compounds including expired 14CO2 within 24 h. The 14C-radioactivity derived from the [14C]sialyl-lactose/sialyl-[3H]lactitol mixture which remained in the bodies of fasted mice after 24 h was less than 1%. In the case of well-fed mice, a higher amount of the sialic acid residues was metabolized. The bulk of radioactivity of the mucin was resorbed within 24 h. About 40% of the radioactivity administered was excreted by the urine within 48 h; 30% of this radioactivity represented sialic acid and 70% other anionic and non-anionic metabolic products. 60% of the radioactivity administered remained in the body, and bound 3H-labeled sialic acids were isolated from liver. Sialyl-alpha (2-3)-[3H]lactitol was injected intravenously into rats; the substance was rapidly excreted in the urine without decomposition. These studies show that part of the sialic acids bound to oligosaccharides and glycoproteins can be hydrolysed in intestine by sialidase and be resorbed. This is followed either by excretion as free sialic acid or by metabolization at variable degrees, which apparently depends on the compound fed and on the retention time in the digestive tract. 相似文献
4.
Studies on the positional integrity of glyceride fatty acids during digestion and absorption in rats 总被引:2,自引:2,他引:0
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1. Rats previously starved for 24hr. were separately given by intraduodenal injections 0.5ml. of a dispersion containing 10mg. of sodium taurocholate, with 50mg. of glycerol 1,3-dioleate 2[1-(14)C]-palmitate, glycerol 1,2-dioleate 3[1-(14)C]-palmitate, a mixture of [1-(14)C]palmitic acid and triolein, or a mixture of [1-(14)C]-palmitic acid and oleic acid. 2. At the end of 30min., the net amounts, and the radioactivity, of the neutral-lipid components recovered from the intestinal lumen and mucosa, and the position of the labelled palmitic acid in the mucosal triglycerides, were determined. 3. When glycerol 1,3-dioleate 2[1-(14)C]-palmitate was administered, most of the labelled acid was retained in the di- and monoglycerides of the lumen; the triglycerides were the major components containing the radioactivity in the mucosa and 75-80% of the labelled acid was located at the beta-position of these triglycerides. 4. When glycerol 1,2-dioleate 3[1-(14)C]-palmitate was administered, the labelled acid was readily split off in the lumen and virtually no radioactivity could be traced in the monoglyceride fraction; in the intestinal mucosa, triglycerides were again the chief components containing most of the radioactivity, and 80-85% of the labelled acid was esterified at the outer positions of the glycerol. 5. When [1-(14)C]palmitic acid mixed with triolein was administered, the concentrations of free fatty acids increased markedly in the intestinal lumen and mucosa, and 80-88% of the radioactivity of the mucosal triglycerides was located at the outer positions of the glycerol. 6. When [1-(14)C]palmitic acid mixed with oleic acid was administered, the labelled acid accumulated in the lumen as well as in the cell, and it was randomly incorporated into all three positions of the mucosal triglycerides. 相似文献
5.
Synthesis of delta-(alpha-aminoadipyl)cysteinylvaline and its role in penicillin biosynthesis. 总被引:12,自引:7,他引:5
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P A Fawcett J J Usher J A Huddleston R C Bleaney J J Nisbet E P Abraham 《The Biochemical journal》1976,157(3):651-660
1. The stereoisomers of delta-(alpha-aminoadipyl)-L-cysteinylvaline (LLD, LLL and DLD) were synthesized from valine labelled with 3H in its methyl groups or in the alpha position. L-Cysteinyl-D-[4,4'-3H]valine was also synthesized. 2. 3H was incorporated into a compound that behaved like penicillin N when the LLD tripeptide containing either a methyl- or an alpha-labelled valine residue was incubated with a cell-free system prepared by lysis of protoplasts of Cephalosporium acremonium. 3. Incorporation was not observed under these conditions from the labelled all-L- or DLD-tripeptide, from L-cysteinyl-D-[4,4'-3H]valine, or of Penicillium chrysogenum appeared to be the LLD isomer, like that from C. acremonium. 5. These findings are discussed in relation to penicillin biosynthesis. 相似文献
6.
M Sugahara 《Journal of biochemistry》1983,93(5):1337-1342
It was found that 4-aminobutyraldehyde (ABAL) is a precursor convertible to gamma-aminobutyric acid (GABA) in vivo. [2,3-3H]ABAL was synthesized from [2,3-3H]putrescine. After the subcutaneous administration of [3H]ABAL at the dose of 1 mumol/g body weight, [3H]GABA was produced in the mouse brain in an amount of about 350 nmol/g brain in 10 min. After oral administration of [3H]ABAL at the dose of 2 mumol/g body weight, [3H]GABA was also produced in the brain in an amount of about 530 nmol/g brain in 30 min. It seems that peripherally administered ABAL penetrates the blood-brain barrier into the central nervous system and is rapidly metabolized to GABA in the brain. 相似文献
7.
Synthesis, characterization and properties of uridine 5′-(α-d-apio-d-furanosyl pyrophosphate) 总被引:1,自引:0,他引:1
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1. A method was developed for synthesizing UDP-apiose [uridine 5'-(alpha-d-apio-d-furanosyl pyrophosphate)] from UDP-glucuronic acid [uridine 5'-(alpha-d-glucopyranosyluronic acid pyrophosphate)] in 62% yield with the enzyme UDP-glucuronic acid cyclase. 2. UDP-apiose had the same mobility as uridine 5'-(alpha-d-xylopyranosyl pyrophosphate) when chromatographed on paper and when subjected to paper electrophoresis at pH5.8. When [(3)H]UDP-[U-(14)C]glucuronic acid was used as the substrate for UDP-glucuronic acid cyclase, the (3)H/(14)C ratio in the reaction product was that expected if d-apiose remained attached to the uridine. In separate experiments doubly labelled reaction product was: (a) hydrolysed at pH2 and 100 degrees C for 15min; (b) degraded at pH8.0 and 100 degrees C for 3min; (c) used as a substrate in the enzymic synthesis of [(14)C]apiin. In each type of experiment the reaction products were isolated and identified and were found to be those expected if [(3)H]UDP-[U-(14)C]apiose was the starting compound. 3. Chemical characterization established that the product containing d-[U-(14)C]apiose and phosphate formed on alkaline degradation of UDP-[U-(14)C]apiose was alpha-d-[U-(14)C]apio-d-furanosyl 1:2-cyclic phosphate. 4. Chemical characterization also established that the product containing d-[U-(14)C]apiose and phosphate formed on acid hydrolysis of alpha-d-[U-(14)C]apio-d-furanosyl 1:2-cyclic phosphate was d-[U-(14)C]apiose 2-phosphate. 5. The half-life periods for the degradation of UDP-[U-(14)C]apiose to alpha-d-[U-(14)C]apio-d-furanosyl 1:2-cyclic phosphate and UMP at pH8.0 and 80 degrees C, at pH8.0 and 25 degrees C and at pH8.0 and 4 degrees C were 31.6s, 97.2min and 16.5h respectively. The half-life period for the hydrolysis of UDP-[U-(14)C]-apiose to d-[U-(14)C]apiose and UDP at pH3.0 and 40 degrees C was 4.67min. After 20 days at pH6.2-6.6 and 4 degrees C, 17% of the starting UDP-[U-(14)C]apiose was degraded to alpha-d-[U-(14)C]apio-d-furanosyl 1:2-cyclic phosphate and UMP and 23% was hydrolysed to d-[U-(14)C]apiose and UDP. After 120 days at pH6.4 and -20 degrees C 2% of the starting UDP-[U-(14)C]apiose was degraded and 4% was hydrolysed. 相似文献
8.
R J Suhadolnik S Pornbanlualap J M Wu D C Baker A K Hebbler 《Archives of biochemistry and biophysics》1989,270(1):363-373
The data presented here describe new findings related to the bioconversion of adenosine to 9-beta-D-arabinofuranosyladenine (ara-A) by Streptomyces antibioticus by in vivo investigations and with a partially purified enzyme. First, in double label in vivo experiments with [2'-18O]- and [U-14C]adenosine, the 18O:14C ratio of the ara-A isolated does not change appreciably, indicating a stereospecific inversion of the C-2' hydroxyl of adenosine to ara-A with retention of the 18O at C-2'. In experiments with [3'-18O]- and [U-14C]-adenosine, [U-14C]ara-A was isolated; however, the 18O at C-3' is below detection. The adenosine isolated from the RNA from both double label experiments has essentially the same ratio of 18O:14C. Second, an enzyme has been isolated and partially purified from extracts of S. antibioticus that catalyzes the conversion of adenosine, but not AMP, ADP, ATP, inosine, guanosine, or D-ribose, to ara-A. In a single label enzyme-catalyzed experiment with [U-14C]adenosine, there was a 9.9% conversion to [U-14C]ara-A; with [2'-3H]-adenosine, there was a 8.9% release of the C-2' tritium from [2'-3H]adenosine which was recovered as 3H2O. Third, the release of 3H as 3H2O from [2'-3H]adenosine was confirmed by incubations of the enzyme with 3H2O and adenosine. Ninety percent of the tritium incorporated into the D-arabinose of the isolated ara-A was in C-2 and 8% was in C-3. The enzyme-catalyzed conversion of adenosine to ara-A occurs without added cofactors, displays saturation kinetics, a pH optimum of 6.8, a Km of 8 X 10(-4) M, and an inhibition by heavy metal cations. The enzyme also catalyzes the stereospecific inversion of the C-2' hydroxyl of the nucleoside antibiotic, tubercidin to form 7-beta-D-arabinofuranosyl-4-aminopyrrolo[2,3-d]pyrimidine. The nucleoside antibiotic, sangivamycin, in which the C-5 hydrogen is replaced with a carboxamide group, is not a substrate. On the basis of the single and double label experiments in vivo and the in vitro enzyme-catalyzed experiments, two mechanisms involving either a 3'-ketonucleoside intermediate or a radical cation are proposed to explain the observed data. 相似文献
9.
The isolation and characterization of 60 nm vesicles (''nanovesicles'') produced during ionophore A23187-induced budding of human erythrocytes. 总被引:3,自引:3,他引:0
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1. Penicillin N was synthesized by coupling alpha-amino-alpha-p-nitrobenzyl-N-p-nitro-benzyloxycarbonyl-D-adipate with 6-aminopenicillanic acid benzyl ester, followed by removal of the protecting groups through hydrogenolysis. 2. alpha-Amino-alpha-p-nitrobenzyl-N-p-nitrobenzyloxycarbonyl-D-[5-14C]adipate was prepared by treating alpha-p-nitrobenzyl-N-p-nitrobenzyloxycarbonyl-D-glutamic acid with [14C]diazomethane followed by rearrangement with silver trifluoromethanesulphonate. 3. Coupling of alpha-amino-alpha-p-nitrobenzyl-N-p-nitrobenzyloxycarbonyl-D-[5-14C]adipate with 6-aminopenicillanic acid benzyl ester gave triprotected [10-14C]penicillin N. 4. 3H was introduced at C-6 of the Schiff's base derivative (10) by oxidation followed by reduction with NaB3H4. 5. The so-derived (6 alpha-3H)-labelled Schiff's base was hydrolysed to give 6-amino [6 alpha-3H]penicillanic acid benzyl ester p-toluenesulphonic acid salt, which after coupling as the free amine with alpha-amino-alpha-p-nitrobenzyl-N-pnitrobenzyloxycarbonyl-D-adipate and then hydrogenolysis, yielded [6alpha-3H]penicillin N. 6. Triprotected [10-14C]penicillin N and triprotected [6alpha-3H]penicillin N in admixture were hydrogenolysed to give [10-14C,6alpha-3H]penicillin N. 相似文献
10.
Metabolism of the diacetyl derivatives of stereoisomeric monoacyl-sn-glycerols by rat adipocytes in vitro.
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Diacetyl long-chain 1(3)- and 2-acyl-sn-glycerols containing either [9,10-3H]oleic acid or [1-14C]palmitic acid were synthesized by partial hydrolysis of the corresponding labelled triacylglycerols and acetylation. They were obtained in a high degree of stereochemical purity by preparative h.p.l.c. on a column containing a diol bonded phase. Each compound was rapidly metabolized by adipocyte preparations in vitro, and a high proportion of the label was recovered in the unesterified fatty acid and triacylglycerol fractions. Negligible amounts of intermediate products of hydrolysis were detected. Triacylglycerols were formed from [9,10-3H]oleic acid and from diacetyl-1(3)-[9,10-3H]oleoyl glycerol precursors at about the same rate, but the 2-isomer was metabolized rather more slowly. The results were consistent with the hypothesis that essentially complete hydrolysis occurred in the medium or at the plasma membrane, through the actions of lipoprotein lipase and monoacylglycerol lipase, and that subsequent esterification took place within the cell. To confirm that no putative intermediate monoacylglycerols were utilized for triacylglycerol biosynthesis via the monacylglycerol pathway, the positional distributions of fatty acids in triacylglycerols from each substrate were determined. No positional selectivity was observed. It was concluded that monoacylglycerols, of an origin exogenous to the tissue, e.g. those derived from plasma triacylglycerols, were not utilized to a significant degree for triacylglycerol biosynthesis in adipose tissue. The diacetyl derivatives of monoacylglycerols may serve as useful stereochemical probes in studies of triacylglycerol biosynthesis via the monoacylglycerol pathway in other tissues. 相似文献
11.
Ehrlich ascites tumor cells were incubated in vitro with rac-1-O-[1'-14C]octadecyl-2-O-methylglycero-3-beta-D-glucopyranosi de for 24 h. The potentially antineoplastic ether glycolipid was rapidly metabolized by the cells to radioactive 1-O-octadecyl-2-O-methylglycerol (70 pmol/10(6) cells per h) and further acylated to 1-O-octadecyl-2-O-methyl-3-acylglycerols. Incubation of Ehrlich ascites cells with synthetic rac-1-O-[1'-14C]octadecyl-2-O-methyl-3-palmitoylglycerol showed that this metabolite is reconverted by deacylation to 14C-labeled 1-O-octadecyl-2-O-methylglycerol. The latter compound or a metabolite derived therefrom may be the 'toxic principle' of both the ether glyceroglycolipids and ether glycerophospholipids having a 1-O-alkyl-2-O-methylglyceryl moiety, as suggested by Unger et al. (J. Natl. Cancer Inst. 78 (1987) 219-222). 相似文献
12.
The removal of partially metabolized very-low-density lipoproteins by the perfused rat liver. 总被引:1,自引:1,他引:0
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1. Donor perfused rat livers were used to prepare VLD (very-low-density) lipoproteins, labelled in their triacylglycerol and protein components with [1-14C]oleic acid and L-[4,5-3H]leucine respectively. Partially metabolized VLD lipoproteins, similarly labelled, were obtained from supradiaphragmatic rats injected with the parent VLD lipoproteins. 2. The triacylglycerol and protein components of the partially metabolized VLD lipoproteins were removed by recipient perfused rat livers at rates much higher than those of the parent VLD lipoproteins. No degradation of the partially metabolized VLD lipoproteins to LD (low-density) lipoproteins occurred during the perfusions. 3. Removal of hepatic lipase from the livers did not significantly affect the rate of removal of the partially metabolized VLD lipoproteins. 相似文献
13.
The metabolic fate of (2-14C)folic acid and a mixture of (2-14C)- and (3'',5'',9-3h)-folic acid in the rat. 总被引:2,自引:2,他引:0
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The metabolism of [2-14C]folic acid over 13 days and a mixture of [2-14C]- and [3',5',9-3h]-folic acid in rats over a 6-day period is described. Both 14C and 3H are excreted in urine over the 6-day period, but 3H and 14C are only detectable in faeces for 2 days. A breakdown product of folic acid labelled with 3H only was found in some urine samples, but no metabolite corresponding to the part of the molecule containing 14C was detected. These experiments show that in the whole animal a substantial portion of orally administered folic acid undergoes scission shortly after administration [Blair Biochem. J. (1957) 68, 385-387] and that the retained folates are a shortage form for folate monoglutamates. 相似文献
14.
A quantitative triterpene analysis was made of latex stem tissue of Euphorbia lathyris. Young plants seedlings of E. lathyris were incubated with various labelled precursors. Incorporation into triterpenes was obtained from [2-14C]mevalonic acid, [1-14C]acetate, [3-14C]pyruvate, [U-14C]sucrose, [U-14C]glucose, [U-14C]xylose, [U-14C]glyoxylate, [2,3-14C]succinic acid, [1-14C]glycerol [U-14C]serine. Both sugars tyrosine appeared to be effective precursors in DOPA synthesis inside the laticifers. Exogenously supplied mevalonic acid was only involved in triterpene synthesis outside the laticifers. GC-RC of triterpenes synthesized from [U-14C]glucose revealed the origin of these compounds in the latex. The labelled triterpenes obtained after incorporation of the other mentioned labelled precursors were only partly synthesized in the laticifers. For quantitative data on latex triterpene synthesis seedlings were incubated with [U-14C]sucrose, [U-14C]glucose, [U-14C]xylose [1-14C]acetate in the presence of increasing amounts of unlabelled substrate. From the amount of 14C incorporated into the triterpenes the amount of substrate directly involved in triterpene synthesis was calculated, as was the absolute triterpene yield. Sucrose showed the highest triterpene yield, equivalent to the daily increase of the triterpene content of growing seedlings. The possible significance of the other precursors in triterpene synthesis in the laticifers is discussed. 相似文献
15.
《Plant science》1987,51(1):97-103
Protoplasts were enzymatically prepared from the mesocarp of two species of oil palm (Elaeis guineensis Jacq. and E. oleifera HBK and Cortes) 16–20 weeks after anthesis and from rapidly multiplying embryogenic cultures of E. guineensis. The protoplasts were purified by density gradient centrifugation in 20% (w/v) sucrose. Radioactive incorporation studies showed that the protoplasts metabolized [1-14C]acetate to lipids, water-soluble compounds and 14CO2. The [14C]fatty acids obtained consisted mainly of C16: 0, C18: 0 and C18: 1. C16: 1, a very minor fatty acid in palm oil, was also labelled and accounted for 8–39% of total fatty acids synthesized by the mesocarp and embryogenic culture protoplasts. The ratio of labelled C18: 0 to C18: 1 was found to vary with the age of the fruit from which the protoplasts were prepared. Thin layer chromatography (TLC) of the labelled lipids showed the presence of all neutral acylglycerol classes. However the distribution of radiolabel in the various classes differed from those previously reported for oil palm mesocarp [K.C. Oo et al. Lipids, 20 (1985) 205] and embryoid tissue slices [E. Turnham and D.H. Northcote, Phytochem., 23 (1984) 35]. Ozonolysis showed that all the labelled C18: 1 acid was vaccenic acid. 相似文献
16.
The conversion of l-[U-(14)C]lysine into carnitine was demonstrated in normal, choline-deficient and lysine-deficient rats. In other experiments in vivo radioactivity from l-[4,5-(3)H]lysine and dl-[6-(14)C]lysine was incorporated into carnitine; however, radioactivity from dl-[1-(14)C]lysine and dl-[2-(14)C]lysine was not incorporated. Administered l-[Me-(14)C]methionine labelled only the 4-N-methyl groups whereas lysine did not label these groups. Therefore lysine must be incorporated into the main carbon chain of carnitine. The methylation of lysine by a methionine source to form 6-N-trimethyl-lysine is postulated as an intermediate step in the biosynthesis of carnitine. Radioactive 4-N-trimethylaminobutyrate (butyrobetaine) was recovered from the urine of lysine-deficient rats injected with [U-(14)C]lysine. This lysine-derived label was incorporated only into the butyrate carbon chain. The specific radioactivity of the trimethylaminobutyrate was 12 times that of carnitine isolated from the urine or carcasses of the same animals. These data further support the idea that the last step in the formation of carnitine from lysine was the hydroxylation of trimethylaminobutyric acid, and are consistent with the following sequence: lysine+methionine --> 6-N-trimethyl-lysine --> --> 4-N-trimethylaminobutyrate --> carnitine. 相似文献
17.
The synthesis of bile acids in perfused rat liver subjected to chronic biliary drainage 总被引:3,自引:2,他引:1
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1. Isolated rat liver was perfused with heparinized whole blood under physiological pressure resulting in the secretion of bile at about the rate observed in vivo. 2. The preparation remained metabolically active for 4h and was apparently normal in function and microscopic appearance. 3. When the perfusate plasma and liver cholesterol pool was labelled by the introduction of [2-(14)C]mevalonic acid the specific radioactivity of the perfusate cholesterol increased. The biliary acids (cholic acid and chenodeoxycholic acid) were labelled and had the same specific radioactivity. 4. Livers removed from rats immediately after, and 40h after, the start of total biliary drainage, were perfused; increased excretion rates of both cholic acid and chenodeoxycholic acid were found when the liver donors had been subjected to biliary drainage. 5. The incorporation of [2-(14)C]mevalonic acid or rat lipoprotein labelled with [(14)C]cholesterol into bile acids was studied. 6. A dissociation between the mass of bile acid excreted and the rate of incorporation of (14)C was found. This was attributed to the changing specific radioactivity of the cholesterol pool acting as the immediate bile acid precursor. 相似文献
18.
Absence of covalently linked core protein from newly synthesized hyaluronate. 总被引:2,自引:1,他引:1
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1. Primary cultures of chondrocytes from the Swarm rat chondrosarcoma were labelled with either [3H]glucosamine or [14C]glucosamine, and hyaluronate synthesized by the cells was isolated from the cell layer. Parallel cultures were labelled with either [3H]serine or [3H]lysine, and identical fractions were isolated from the cell layer. Some cultures were dual-labelled. 2. In cultures labelled with [3H]serine for between 30 min and 24 h and extracted with 4.0 M-guanidine, a procedure that solubilizes predominantly extracellular macromolecules, small amounts of [3H]serine-labelled molecules were found associated with the hyaluronate fraction purified from the extract by dissociative CsCl-density-gradient centrifugation and dissociative Sepharose CL-2B chromatography. About 75% of the [3H]serine-labelled molecules in the fraction were specifically associated with hyaluronate, since they could be removed by prior treatment with proteinase-free Streptomyces hyaluronidase. The association of the [3H]serine-labelled molecules with hyaluronate was non-covalent, since they could be separated from it by further centrifugation in CsCl density gradients containing 4 M-guanidinium chloride and a zwitterionic detergent. 3. In other experiments the cultures were extracted with a sequential zwitterionic-detergent/guanidinium chloride procedure that completely solubilized the cell layer and enabled fractions containing newly synthesized cell-associated hyaluronate to be isolated. Zwitterionic detergent was present throughout. No [3H]lysine was incorporated into these fractions, irrespective of whether the cultures were pulsed concurrently with [3H]lysine and [14C]glucosamine or sequentially with [3H]lysine to prelabel the protein pool (24 h) followed by [14C]-glucosamine to label hyaluronate (1 h). 4. The results show that newly synthesized hyaluronate is not associated with covalently bound protein, and suggest that chain synthesis is initiated by a mechanism other than on to a core protein. Small amounts of [3H]serine-labelled molecules are, however, non-covalently associated with extracellular hyaluronate. Their identity is at present unknown, but they are probably of low molecular weight. 相似文献
19.
1. [1-(14)C]Ethyl carbamate, ethyl [carboxy-(14)C]carbamate, [1-(14)C]ethanol and sodium hydrogen [(14)C]carbonate were injected intraperitoneally into C57 mice, and nucleic acids and proteins were separated from the liver and lungs with phenol as described by Kirby (1956). 2. Chromatographic analysis of the hydrolytic products of the urethane-labelled RNA showed the presence of a single radioactive compound differing in behaviour from the major pyrimidine nucleotides and purines. 3. The products from RNA labelled by [1-(14)C]ethyl carbamate or ethyl [carboxy-(14)C]carbamate appeared chromatographically identical but could not be detected in the RNA of mice given [1-(14)C]ethanol or sodium hydrogen [(14)C]-carbonate. 4. The labelled product appeared to be the ethyl ester of cytosine-5-carboxylic acid formed by the reaction of urethane with RNA in vivo. 5. A direct reaction between labelled urethane or the labelled metabolite of urethane, [1-(3)H]-ethyl N-hydroxycarbamate, and RNA was not detected. 相似文献
20.
H H Schmid P C Bandi N C Chang T H Madson W J Baumann 《Biochimica et biophysica acta》1975,409(3):311-319
1-O-[1'-14C]Hexadecyl ethanediol was administered intracerebrally to myelinating rat brain, and incorporation of radioactivity into brain lipids was followed over a 48-h period: (1) O-Hexadecyl ethanediol was metabolized primarily through oxidative ether bond cleavage, and much of the label was recovered in phospholipid acyl groups. (2) Substantial amounts of radioactivity were also found in choline and ethanolamine phospholipids having an O-hexadecyloxyethyl glycerol backbone. This means that alkyl ethanediol was used in glycerol ether biosynthesis as are long-chain primary alcohols. (3) Acidic hydrolysis of the ethanolamine glycerophosphatide fraction yielded also labeled hexadecanol which may indicate desaturation of 1-O-hexadecyloxyethyl 2-acyl glycerophosphoryl ethanolamine to the plasmalogen analogue. (4) Small amounts of the substrate were oxidized to O-hexadecyl glycolic acid and incorporated into the phospholipids. The substrate did not serve as precursor of O-hexadecyl ethanediol phosphorylcholine or phosphorylethanolamine in the brain. 相似文献