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1.
A method is presented for purifying papain from extracts of papaya latex. The procedure involves precipitation of the extract of papaya latex with sodium chloride followed by affinity chromatography of the redissolved precipitate. Precipitation of the protein from the latex extract is necessary to separate the papain from material which interferes with the binding of papain to the affinity column. During affinity chromatography, the affinity column is overloaded to insure absence in the final product of impurities which are capable of binding to the affinity column.The papain prepared by this procedure yielded an amino acid analysis and an N-terminal amino acid analysis expected for a sample of pure papain. No Met was detected on amino acid analysis nor was the presence of N-terminal residues other than He detected. On polyacrylamide disc gel electrophoresis at pH 4.3, papain prepared by the method described in this work was indistinguishable from crystalline papain which was prepared by the method of Kimmel and Smith, and further purified by affinity chromatography. Both disc gel patterns consisted of a single band and a trailing shadow which was less than 5% of the main band. In routine spectrophotometric assays, the specific activity toward N,α-benzoyl-l-arginine ethyl ester of papain prepared by the procedure described in this work was indistinguishable from crystalline papain prepared by the method of Kimmel and Smith, and further purified by affinity chromatography. Values of 24 sec?1' and 15 mm were obtained from the turnover number and Km for the papain-catalyzed hydrolysis of N,α-benzoyl-l-arginine ethyl ester at 25 °C, pH 6.00, using a pH stat. 相似文献
2.
The and metabolism of 1,2- 3H-progesterone was studied in estrogen-stimulated and control vaginae of ovariectomized mice. Employing two-dimensional thin-layer chromatography, gas-liquid chromatography and metabolite “trapping” techniques, the major and minor pathways for progesterone metabolism were determined and shown to involve saturation of the Δ4-double bond to yield 5α-pregnane compounds and reduction of the C20 and C3 ketone groups to form 20α- and 3α- and 3β-hydroxy derivatives, respectively. The quantities of 20β-hydroxy metabolites and 5β-epimers that were detected were considered not to be significant. The major metabolites formed by untreated tissues following incubation in the presence of both high (10?6M) and low (10?8M) progesterone concentrations were 3α-hydroxy-5α-pregnan-20-one and 5α-pregnane-3,20-dione. Although these two derivatives were also found in sizable quantities in estrogen-treated tissues, a marked increase (5-fold) in the rate of C20 ketone reduction at high progesterone concentrations (10?6M) to yield 20α-hydroxy-4-pregnen-3-one was demonstrated. Following intravaginal administration of 3H-progesterone , only progesterone and 3α-hydroxy-5α-pregnan-20-one were retained in appreciable quantities through 2 hr, suggesting rapid loss of 20α-hydroxy-4-pregnen-3-one and the 5α-pregnanediols from this tissue under conditions. 相似文献
3.
Bojan Hamlin Jennings Sharon Martin Boudreau Beverly Braasch Cheryl Garganta 《Steroids》1982,39(4):371-380
Both the 5α, 6α- and 5β, 6β-dichloromethylene adducts ( and ) of 3β-acetoxy-5-androsten-17-one () are produced when the latter is exposed to dichlorocarbene generated from chloroform and base by Phase Transfer Catalysis using ultrasound as a means of agitation. The 1H NMR substituent effects of 5α, 6α- and 5β, 6β-dichloromethylene on the angular methyl groups (Zürcher values) are given. The 13C NMR spectra for both compounds are presented and discussed. 相似文献
4.
The frequency of spontaneous contractions of seminiferous tubules of the rat appeared to be increased in a dose-dependent manner by prostaglandin F1α. PGF1α treatment increased the tonus of the smooth muscle cells in the wall of the tubules as indicated by a reduction in the diameter of the tubules. When the tubules were rinsed successively with fresh Tyrode's solution, the contractile frequency was diminished. Returning the original bathing medium to the tubules restored their contractile frequency, as did treatment of the rinsed tubules with PGF1α (10-7 M). Pre-injecting the rats with indomethacin tended to reduce the contractile frequency of the extirpated tubules. Treating the tubules with a solution of indomethacin for 90 min. was more effective than pretreatment in reducing contractile frequency, but a combination of these two procedures produced the greatest inhibition. PGF1α restored the contractile frequency of the indomethacin-treated tubules. Our results indicate that PGs modulate the contractility of the tubules. 相似文献
5.
Mohammad Ali Ghalambor Christopher D. Warren Roger W. Jeanloz 《Biochemical and biophysical research communications》1974,56(2):407-414
Incubation of UDP-[14C]--acetylglucosamine with calf pancreas microsomes in the presence of Mn++ and potassium thiocyanate gave a labeled glycolipid, tentatively identified as 1-2-acetamido-2-deoxy--glucosyl 2-dolichyl pyrophosphate on the basis of cochromatography with synthetic 1-2-acetamido-2-deoxy-α--glucopyranosyl 2-dolichyl pyrophosphate, similar chemical and enzymic hydrolyses of the biosynthetic and synthetic compounds, and stimulation of the biosynthesis by addition to the incubation mixture o dolichyl phosphate or a crude lipid fraction extracted from microsomes. 相似文献
6.
Göran Hansson 《Prostaglandins & other lipid mediators》1979,18(5):745-771
The metabolism of the prostaglandin F2α analogues, 15-methyl-Δ4--PGF1α and 16,16-dimethyl-Δ4--PGF1α, has been investigated in the cynomolgus monkey and the human female. The two analogues, tritium labelled in the 9β-position, were administered by intramuscular injections into the monkeys and by subcutaneous injections into the human. Excretion of tritium labelled products were followed in urine (in both species) and feces (in monkeys only) and several metabolites were identified by GC/MS. The analogues were found to be resistant to the 15-hydroxy dehydrogenase and furthermore the degradation by β-oxidation was delayed. About 13% of the given dose of 15-methyl-Δ4--PGF1α was excreted unchanged into urine and feces from the monkey. The corresponding figure for 16,16-dimethyl-Δ4--PGF1α was about 20%. In addition, a large part of the metabolites had the carbon skeleton intact and were only metabolized by ω-oxidation. The relative resistance to degradation of these two analogues is likely to be the basis for their prolonged pharmacological activity. 相似文献
7.
Two synthetic routes to 3-deoxy-1α-hydroxyvitamin D3, an analog of 1α,25-dihydroxyvitamin D3, are described. One involved the six-step conversion of 1α,2α-epoxy-6,6-ethylenedioxy-5α-cholestan-3- one to 1α-acetoxycholest-5-ene, whereas, in the second, the same intermediate was prepared from 1α-hydroxycholesterol. Conversion of the Δ5-sterol to the required 5,7-diene was accomplished most efficiently via 7-keto and 7-tosylhydrazone intermediates. Bioassay of 3-deoxy-1α-hydroxyvitamin D3 in the rat establishes that the analog can fulfill all common vitamin D functions including stimulation of intestinal calcium transport, mobilization of calcium and phosphate from bone, stimulation of growth, and calcification of bone. Direct comparison indicates the compound to have to of the activity of 1α-hydroxyvitamin D3, but it acts with a time course indistinguishable from the latter. 相似文献
8.
A series of 12α-hydroxy steroids with varying side chains was prepared, and their 24-hour acetylation yields were compared, l2α-Hydroxy-5β-pregnan-20-one () was prepared from 3α, 12α-diacetoxy-5β~pregnan-20-one () and also by side chain degradation of 12α-acetoxy-5β-cholanoic acid (). 21-Benzyl-5β-pregnan-12α-ol () was synthesized by hydrogenation of the 21-benzylidine derivative of ketone . 23-Pheny1-5β-norcholan-12α-ol () was obtained by the Grignard reaction of 2-phenyl-ethylmagnesium bromide and ketone , dehydration, hydrogenation and hydride reduction; a similar sequence produced 20-methyl-5β-pregnan-12α-ol (). The acetylation results (Table 11) imply that branching at C-20 may be more significant for 12α-hydroxyl reactivity than side chain length or type. An additional compound with an unbranched side chain, 21-nor-5β-cholan-12α-ol (), was synthesized by a Grignard reaction on the 21-bromo intermediate . Acetylation rates determined by glc indicate (Table 111) That compounds with unbranched side chains have 12α-hydroxyl groups about ten times as reactive as their analogs with 20-methyl groups. 相似文献
9.
William Egan Fai-Po Tsui Peggy Ann Climenson Rachel Schneerson 《Carbohydrate research》1980,80(2):305-316
The capsular polysaccharide from Klebsiella K44 has been investigated by the techniques of methylation, base-catalyzed elimination, Smith degradation, and partial hydrolysis. The last-named yielded an oligosaccharide corresponding to one repeating unit. The anomeric configutations of the sugar residueswere determined by 1H- and 13C-n.m.r. spectroscopy. The polysaccharide has a fractional acetyl content and is the first in this series to be based on a linear, pentasaccharide repeating unit. 相似文献
10.
《International journal of biological macromolecules》1987,9(5):308-310
(d.s. 0.86 for trityl) was oxidized with dimethyl sulphoxide-phosphorus pentoxide in to give an oxidized product, which was converted to its oxime (d.s. 0.91 for oxime). Hydrogenation of the oxime with lithium aluminium hydride in tetrahydrofuran, followed by detritylation with hydrochloric acid in methanol gave an aminodeoxycellulose (d.s. 0.5 for amino group) in a 46% yield. 2-Amino-2-deoxy-d-glucose and 2-amino-2-deoxy-d-mannose, d-glucose and d-mannose (4:1:4:1), which were detected by g.l.c. and p.c. in the acid hydrolysate of the acetamidodeoxy derivative. Neither the acetamidodeoxy nor aminodeoxy celluloses were hydrolysed by chitinase from Streptomyces griseus and lysozyme from hen egg-white. 相似文献
11.
D.H. van den Eijnden D.H. Joziasse L. Dorland H. van Halbeek J.F.G. Vliegenthart K. Schmid 《Biochemical and biophysical research communications》1980,92(3):839-845
Partial sialylation of biantennary and triantennary glycopeptides of α1-acid glycoprotein using colostrum β-galactosideα(2→6) sialyltransferase followed by high resolution 1H-NMR spectroscopic analysis of the isolated products enabled the assignment of the branch as the most preferred substrate site for sialic acid attachment. The branch appeared to be much less preferred and the Galβ(1→4)GlcNAcβ(1→4)Manα(1→3)Man sequence of triantennary structures was of intermediate preference for the sialyltransferase. The specificity of the β-galactoside α(2→6) sialyltransferase is thus shown to extend to structural features beyond the terminal -acetyllactosamine units on the oligosaccharide chains of serum glycoproteins. 相似文献
12.
Assay of liver microsomal cholesterol 7 alpha-hydroxylase using deuterated carrier and gas chromatography-mass spectrometry 总被引:1,自引:0,他引:1
The activity of cholesterol 7α-hydroxylase in rat liver microsomes was assayed by measuring the mass of 5-cholestene-3β, 7α-diol formed from endogenous cholesterol under standardized incubation conditions. After termination of incubations, a known amount of 5-[24,25,7β-2H3]cholestene-3β,7α-diol was added. A chloroform extract of the incubation mixture was subjected to thin layer chromatography and the fraction containing 5-cholestene-3β,7α-diol was converted into trimethylsilyl ether. The trimethylsilyl ether was subjected to combined gas chromatography-mass spectrometry and the amount of unlabeled 5-cholestene-3β,7α-diol in the mixture was calculated from the ratio between the relative intensitics of the peaks at 456 (M-90) and 459 [M-(90 + 3)]. The precision of the method was ±2.2% (SD). The results with this method of assay of cholesterol 7α-hydroxylase were compared with those obtained with a method based on conversion of a trace amount of added [4-14C]cholesterol into 5-cholestene-3β,7α-diol. 相似文献
13.
Zoltán Spolarics Jozsef Mandl Raymund Machovich Patrick Lambin Tamás Garzó Ferenc Antoni István Horváth 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1985,845(3):389-395
125I-labelled complexed with thrombin or plasmin bound to hepatocytes in a concentration-and time-dependent manner. The apparent values calculated from displacement experiments were 7.9 · 10?8 M for and 8.5 · 10?8 M for . Association of these complexes was only partially reversible; after a 180 min incubation period, 50–60% of the bound radioactivity was internalized by the cells. itself bound also to hepatocytes, but the affinity of the complexes was higher than that of the inhibitor alone, and was not internalized, either. 125I-labelled thrombin or plasmin bound to hepatocytes as well. These bindings were also concentration-dependent and could be decreased with an excess of unlabelled ligands. Binding rates and amounts of the bound proteinases were higher than those of their complexes. The complex competed with the complex in binding to hepatocytes, whereas there was no competition between these complexes and the antithrombin III-thrombin complex. These results suggest that the binding sites of hepatocytes for and antithrombin III-proteinase complexes are different. 相似文献
14.
T Shiratori 《Life sciences》1974,14(5):929-935
Tritium- -tocopherol was incorporated into chylomicra by feeding dl-α-(5 methyl 3H)-tocopherol to rats with thoracic duct cannulae. Tritium α-tocopherol in chylomicra, isolated by flotation through 0.85% NaCl, was injected intravenously into rats and distribution of radioactivity was followed in skin, viscera and carcass. Label measured at following injection demonstrated a rapid uptake and prolonged retention in skin compared to viscera and carcass. In fact, relative distribution in skin increased from an average of 10.3% at to 40.1% of the recovered dose at 20 days. Also, the uptake of isotope by skin, expressed as 3H dpm/g lipid, markedly exceeded that of epididymal fat suggesting a preferential distribution to cutaneous tissue. Significant radioactivity was found on the skin surface and hair following injection of 3H-α-tocopherol demonstrating the excretion or secretion of vitamin E compound(s) by the skin. 相似文献
15.
Susumu Sunaga Noboru Horiuchi Naoyuki Takahashi Kazuo Okuyama Tatsuo Suda 《Biochemical and biophysical research communications》1979,90(3):948-955
Metabolism of 25-hydroxyvitamin D3 (25-OH-D3) in pregnancy was investigated in New Zealand White rabbits fed a rabbit chow. Kidney homogenates from pregnant mothers and fetuses were separately incubated with [3H]-25-OH-D3. The homogenates from fetuses produced significant amounts of D3 [1α,25-(OH)2-D3] from its precursor, while those from mothers predominantly produced [3H]-24,25-dihydroxyvitamin D3 [24,25-(OH)2-D3]. The identity of the radioactive metabolites produced from [3H]-25-OH-D3 was established by periodate cleavage and comigration with synthetic 1α,25-(OH)2-D3 or 24,25-(OH)2-D3 on high pressure liquid chromatography. These results clearly indicate that the fetal kidney is at least one of the sites of 1α,25-(OH)2-D3 synthesis in pregnancy. 相似文献
16.
The pattern of androgenic metabolites in blood, muscle, caput and cauda epididymidis has been investigated in functionally hepatectomized 24 hours castrated rats, 3 hours after the intra-muscular injection of 200 μCi of 3H -3α-diol. Identification of the radioactive metabolites showed only negligible differences between the epididymal regions. In both caput and cauda the main metabolite was DHT (17β-hydroxy-5α-androstane-3-one); 3α- and 3β-diol, androsterone (3α-hydroxy-5α-androstane-17-one), 5-A-dione (5α-androstane-3,17-dione), Δ16-3α-ol (5α-androst-l6-en-3α-ol), Δ16-3β-ol (5α-androst-l6-en-3α-ol) and Δ16-3-one (5α-androst-l6-en-3-one) were also present. and were the predominant metabolites in blood and muscle. No Δ16 compounds could be detected and in constrast to epididymis, more than 50% of the radioactivity was associated with polar compounds. From determination of total radioactivity, it was seen that retention by epididymis varied from two to four times that of muscle. Purification and identification of the radioactivity associated with the nuclear fraction demonstrated that DHT was the only nuclear bound androgen.It is suggested from these results that at least one effect of 3α-diol on the rat epididymis is exerted through its conversion to DHT. 相似文献
17.
Satoshi Ōmura Hiroie Ohno Tsuneo Saheki Masakazu Yoshida Akira Nakagawa 《Biochemical and biophysical research communications》1978,83(2):704-709
Elasnin, a new human granulocyte elastase inhibitor, has been isolated from KM-2753. Elasnin is a neutral, lipophilic colorless and viscous oil (, , λmaxEtOH 291 nm (ε, 7760)). The molecular formula was C24H40O4 (M.W.: 392) as determined by its elemental analysis and mass spectrum. Elasnin inhibits markedly human granulocyte elastase, but is almost ineffective for pancreatic elastase, trypsin, chymotrypsin, thermolysin and papain. 相似文献
18.
The epididymis of adult rats metabolize 3H-testosterone by experiments . Thirty minutes after the injection of 100 μCi 3H-testosterone, some 10 per cent of the total radioactivity of the epididymis was found in the water-soluble fraction, whereas 90 per cent was found in the ether soluble fraction (free steroids). The free steroids were examined further and the following androgenic metabolites identified: (17β-hydroxy-4-androsten-3-one) 8, 9%, (4-androstene-3, 17-dione, 2,7%, (5α-androstane-3, 17-dione) 6,5%, (17β-hydroxy-5α-androstan-3-one) 47, 2%, (5α-androstane-3β, 17β-diol) 4, 4%, (5α-androstane-3α,17β-diol) 20, 8% and (3α-hydroxy-5α-androstan-3-one) 3,4%. The relative amount of each metabolite is given in per cent of total radioactivity in the ether soluble fraction. 相似文献
19.
E C Trout 《Steroids》1974,23(1):133-144
The effects of age and sex on the induction by 3,5,3′-triiodothyronine (T-3) of cortisone Δ4-5α-reductase and glucose 6-phosphate dehydrogenase (G 6-P D) in liver and the latter in adrenal have been investigated. Levels of cortisone Δ4-(5α, 5β)-reductase and G 6-P D were measured in homogenates of tissue from normal and T-3 injected male and female rats, to 21 months of age. Increases in the levels of the reductase seen under T-3 stimulation were ascribed to induction of the 5α-reductase alone. T-3 caused induction of cortisone Δ4-5α-reductase only in the livers of male rats months of age. There was induction of total G 6-P D at most ages except in the livers of old male and young female rats and adrenals of young and old male rats. At all ages in normal animals of both sexes the maximum activity of glucose 6-phosphate dehydrogenase was much greater than that of cortisone Δ4-(5α, 5β)-reductase. It is concluded that the amount of G 6-P D in normal liver may be sufficient to handle an increase in cortisone reduction, and factors other than cortisone Δ4-reductase or G 6-P D levels alone must regulate increased reduction of the steroid. 相似文献
20.
Following incubation with [3H]-PGF2α, 73–91% of the 3H activity accumulated by rabbit uterus, choroid plexus or anterior uvea was shown to remain associated with PGF2α on two different chromatographic systems. The tissue to medium ratios, calculated on the basis of chromatographically identified [3H]-PGF2α, were greater than unity (2.3–10.4) for all three tissues and the extracted 3H activity could be effectively accumulated by these tissues for a second time. Under conditions when 85% of authentic [3H]-PGF2α and only 8% of [3H]-15-keto-PGF2α was adsorbed on rabbit anti-PGF2α serum, 60–75% of the extracted 3H was adsorbed onto the antiserum. Following incubation with a mixture of 5,6-[3H]-PGE1 and 2-[14C]-PGE1, the anterior uvea and the uterus showed similar ratios for 3H and 14C and the ratios were essentially constant in their respective homogenates, extracts and chromatographic fractions, indicating insignificant β-oxidation of the accumulated PGE1. In the case of the kidney cortex, a substantial fraction of the accumulated 14C did not extract as a PG presumably as a result of β-oxidation. It is concluded that metabolic alteration of the accumulated PG molecule does occur in some tissues, but such chemical alterations are not an integral part of the PG accumulative process. These results are consistent with the concept that some vertebrate tissues can accumulate PGs against a concentration gradient by an active transport mechanism. 相似文献