Sodium dodecyl sulfate, concurrent inhibitor of several dehydrogenases

The effect of sodium dodecyl sulfate on the activity of highly purified or crystalline enzymes has been studied. The enzymes were: lactate dehydrogenase (LDH), malate dehydrogenase (MDH). isocitrate dehydrogenase (ICDH), glucose-6-phosphate dehydrogenase (G6P-DH), lipase, alkaline phosphatase. Sodium dodecyl sulfate, always under the critical micellar concentration, shows a selective inhibitory effect. A kinetic analysis of the inhibitory action on LDH, MDH, ICDH and G6P-DH was also carried out.     

The effect of bile salts on human vascular endothelial cells

The uptake and release of radiochromium from adult human vascular endothelial cells in culture was employed to determine the relative toxicity of different bile salts. Endothelial cells after pre-incubation with 51Cr for 18 h were incubated with bile salts for 24 h and percentage chromium release was taken as a measure of toxicity to cells. Lithocholic acid (LC) (potassium salt) was cytotoxic at concentrations greater than 50 microM. However, LC glucuronide, sulfate and the beta-epimer were progressively less toxic with toxicity seen at concentrations of 60, 110 and 180 microM, respectively. The greatest cytotoxic effect was observed with glycolithocholic acid (GLC) (potassium salt) which was toxic at every concentration tested (20-200 microM). Sulfation abolished the toxic effect of GLC. At the concentrations employed for the assay (between 20 and 240 microM) GLC sulfate (disodium salt), taurolithocholic acid sulfate (disodium salt), cholic acid (sodium salt), glycocholic acid (sodium salt), deoxycholic acid (sodium salt) and ursodeoxycholic acid (sodium salt) were not cytotoxic. The 51Cr release cytotoxicity assay was validated with lactate dehydrogenase leakage from endothelial cells with a good correlation (r = 0.87). These data confirm in a human cellular system that LC and its conjugates were the most toxic of the bile salts tested and explains its pathophysiological importance in hepatobiliary disease. It also suggests that biotransformation by either sulfation or beta-epimerisation of bile salts especially of LC, as occurs in patients with intrahepatic or extrahepatic biliary obstruction or severe cholestasis, is hepatoprotective.     

The palmitoleate: a natural selective denaturant of enzymes

A study has been carried out in order to explain the enzyme-palmitoleate interaction. The highly purified and crystalline enzymes representative of fundamental metabolic pathways were: alcohol dehydrogenase (ADH), lactate dehydrogenase (LDH), malate dehydrogenase (MDH), isocitrate dehydrogenase (ICDH), glucose-6-phosphate dehydrogenase (G6P-DH), alkaline phosphatase. The enzyme-palmitoleate interaction was studied as a phenomenon time-independent (inhibition) and time-dependent (inactivation). Palmitoleate inhibited remarkably LDH, MDH, ICDH and G6P-DH. A kinetic analysis of the inhibitory action of palmitoleate on LDH and MDH was also carried out. Inactivation studies have shown that ADH and alkaline phosphatase are not sensitive to palmitoleate action, unlike the other enzymes. A comparison was made between the action of palmitoleate and that of a synthetic anionic detergent, sodium dodecyl sulfate (SDS).     

Studies on scavenger receptor inhibitors. Part 1: synthesis and structure-activity relationships of novel derivatives of sulfatides

Scavenger receptors have been proven to be implicated in the formation of atherosclerotic lesions. A series of novel derivatives of sulfatides were synthesized, and their inhibitory activities against incorporation of DiI-acetyl-LDL into macrophages were evaluated in order to clarify the structure-activity relationships of sulfatides as a scavenger receptor inhibitor and find out novel inhibitors with synthetic easiness. The chemical modification of the substructures of sulfatides led to the establishment of the following structure-activity relationships; (1) the ceramide moiety can be replaced with another structure bearing two long chains, (2) the galactose moiety can be replaced with another structure or be deleted without a large decrease in the inhibitory activity, (3) the sulfate moiety was crucial, and it was the most preferable functional group for a potent inhibitory activity. The inhibitory activity of (S)-2-octadecanoylamino-2-tetradecylcarbamoyl)ethyl sulfate sodium salt (3a) against incorporation of DiI-acetyl-LDL into macrophages was proven to be based on the inhibition against the binding of acetyl-LDL to the surface of macrophages. We discovered novel scavenger receptor inhibitors with synthetic easiness, such as (S)-2-octadecanoylamino-2-(tetradecylcarbamoyl)ethyl sulfate sodium salt (3a) and 2-octadecanoylamino-1-(octadecanoylaminomethyl)ethyl sulfate sodium salt (13q).     

Salt Tolerance in Suaeda maritima (L.) Dum: THE EFFECT OF SODIUM CHLORIDE ON GROWTH, RESPIRATION, AND SOLUBLE ENZYMES IN A COMPARATIVE STUDY WITH PISUM SATIVUM L

The effect of sodium, chloride on the growth of a halophyte,Suaeda maritima (L.) Dum., was compared with its effect on Pisumsativum L. cv. Alaska under controlled environmental conditions.The salt stimulated the growth of Suaeda maximally at concentrationsof 170 to 340 mM while the growth of Pisum was inhibited evenby 100 mM. Both species accumulated ions in the tops and themaximum concentrations of Na⁺ and Cl^– rose in Suaeda to860 mM (based on the water content) and 730 mM and in Pisumto 170 mM and 300 mM respectively. Respiration in both specieswas inhibited as the NaCl level in the culture solution wasraised. Four supernatant enzymes (malic dehydrogenase, glucose-6-phosphatedehydrogenase, peroxidase, and acid phosphatase) prepared fromPisum and from Suaeda (grown either in the absence of addedNaCl or in the presence of 340 mM NaCl) were assayed in variouslevels of sodium chloride. The dehydrogenases were markedlyinhibited by increasing salt concentrations while there wasa smaller effect on the peroxidase and acid phosphatase. Therewas no difference in the effect of salt on the enzymes preparedfrom the two species although one is halophilic and the otherhalophobic.     

Properties of a halophil nicotinamide–adenine dinucleotide phosphate-specific isocitrate dehydrogenase. Preliminary studies of the salt relations and kinetics of the crude enzyme

The effects of chlorides on NADP-specific isocitrate dehydrogenase from Halobacterium salinarium were investigated. The enzyme is stabilized by potassium chloride and sodium chloride and this effect is discussed in relation to the Hill (1913) equation. Kinetics of the enzyme were studied within a range of concentrations of potassium chloride and sodium chloride. Apparent Michaelis constants for both substrates were affected by salt concentration, the effect being greater in sodium chloride than in potassium chloride. Minimal apparent Michaelis constants for both substrates were similar to the corresponding constants reported for yeast isocitrate dehydrogenase. V(max.) was maximal in each salt at a concentration of about 1m. The maximum was higher in sodium chloride than in potassium chloride. At salt concentrations above about 2.3m, the apparent V(max.) was lower in sodium chloride than in potassium chloride, and at salt concentrations below 0.75-1.0m, each salt behaved as a linear activator of the enzyme. Within this concentration range salt and NADP(+) acted competitively; the activation by salt was overcome at finite concentrations of NADP(+). At concentrations above about 1m, potassium chloride was a linear non-competitive inhibitor of the enzyme. Within the range 1.0-2.5m, sodium chloride was also a linear non-competitive inhibitor, but above 2.5m it caused more pronounced inhibition.     

Modelling the behaviour of Listeria monocytogenes in ground pork as a function of pH, water activity, nature and concentration of organic acid salts

AIMS: to study and model the effect of sodium acetate, sodium lactate, potassium sorbate and combination of acid salts on the behaviour of Listeria monocytogenes in ground pork. METHODS AND RESULTS: Water activity (a(w)), pH and concentration of acid salt of the meat were adjusted. The behaviour of inoculated L. monocytogenes was studied and modelled according to physicochemical parameters values. Whatever the acid salt concentration used, we observed an inhibition of the growth of L. monocytogenes at pH 5.6 and a(w) 0.95. At pH 6.2 and a(w) 0.97, addition of 402 mmol l(-1) of sodium lactate or 60 mmol l(-1) of potassium sorbate was required to observe a slower growth. CONCLUSIONS: The inhibitory effect of acid salts was a function of pH, a(w), as well as of the nature and concentration of acid salts added. When one acid salt was added, the Augustin's model (Augustin et al. 2005) yielded generally correct predictions of either the survival or growth of L. monocytogenes. SIGNIFICANCE AND IMPACT OF THE STUDY: The suggested model can be used for risk assessment concerning L. monocytogenes in pork products.     

Response of the rat's hepatic drug-metabolizing enzyme system to chemotherapy of Trypanosoma b. brucei infections with Berenil and Suramin

Berenil (4,4-diamidinodiazoaminobenzene-diacetamide acetate) or Suramin [sodium salt of 8-(3-benzamido-4-methylbenzamido)-naphthalene-1,3,5-trisulfonic acid] treatment of rats infected with Trypanosoma b. brucei enhanced hepatic microsomal aniline hydroxylase and p-aminopyrine N-demethylase activities. While Suramin inhibited significantly the activities of cytoplasmic glutamate dehydrogenase and lactate dehydrogenase, Berenil had no effect. The kinetic profiles of these enzymes consistently showed a Km value similar to that of controls. Both cytosolic and microsomal glutathione-S transferase and microsomal epoxide hydratase were unaffected by Suramin. However, a significant increase in cytosolic glutathione-S transferase was observed with Berenil. Microsomal phospholipids were not affected by any of the drugs.     

化学诱导龙血树产生血竭的初步研究

血竭是一种名贵的传统中药,疗效显著,资源稀缺。人工种植龙血树并诱导龙血树产生血竭是解决血竭原料短缺的根本途径。以柬埔寨龙血树为材料,采用26种具有诱导其他植物增加抗性能力的化合物分别接入龙血树进行诱导生产血竭。结果表明:草酸、赤霉素、硫酸锌、芸苔素内酯、硝酸镁、5-硝基愈创木酚酸钠、2-硝基苯酚钠、盐酸、硫代硫酸钠、白氨酸、激动素、IAM、草酸钠、4-硝基苯酚钠、复硝酚钠和水杨酸等16种化合物能增产血竭;增产率达到100%以上的化合物有:1%的硫酸锌,1%的草酸,5 mg·L-1的赤霉素,0.1%硝酸镁,0.1%4-硝基苯酚钠;1%的草酸增产率214%,为最高,其次是5 mg·L-1的赤霉素,增产率为157%;生长素和赤霉素混合使用增产率为183%。     

Purification and biochemical characterization of tellurite-reducing activities from Thermus thermophilus HB8.

Cell-free extracts of Thermus thermophilus HB8 catalyze the in vitro, NADH-dependent reduction of potassium tellurite (K2TeO3). Three different protein fractions with tellurite-reducing activities were identified. Two exhibited high molecular weight and were composed of at least two different polypeptides. The protein in the third fraction was purified to homogeneity and had a single polypeptide chain of 53 to 54 kilodaltons, with an isoelectric point of 8.1. Each enzyme was thermostable, the temperature optimum was 75 degrees C, and 30 mM NaCl, 1.5 M urea, or 0.004% sodium dodecyl sulfate caused 50% inhibition of the enzymes. However, 2% Triton X-100 did not have an inhibitory effect. The enzymes were also able to catalyze the reduction of sodium selenite and sodium sulfite in vitro. NADH was replaceable by NADPH. Divalent cations, such as Ca2+ and Ba2+, had no effect on the activity, while similar concentrations of Zn2+, Ni2+, and Cu2+ abolished the activity. This reductase activity could enable these bacteria both to reduce K2TeO3 and to increase their tolerance toward this salt.     

Effect of different salts and their concentration on polymyxin M biosynthesis by Bac. polymyxa Ross]

The effect of various concentrations of ammonium sulfate, sodium and potassium chlorides and sulfates on biosynthesis of polymyxin M by Bac. polymyxa Ross. on media containing wheet meal as the carbon source was studied. The culture growth, antibiotic accumulation and consumption of the main nutrients was studied as dependent on the level of the salts in the medium. The highest productivity was observed on media with the salt concentrations amounting to 1--2 per cent except sodium sulfate: changes in the concentration of the latter had practically no effect on the activity of the fermentation broth.     

CO2／盐冲击对小麦幼苗呼吸酶活性的影响

以不同抗盐小麦 (Triticum aestivum L.)为材料 ,研究了 CO2 /盐冲击对幼苗生长状况、叶绿素含量、光呼吸和三羧酸循环 (TCAC)关键酶活性的影响。结果表明 :Na Cl抑制小麦生长 ,而 CO2 促进生长 ,这种效应盐处理植株比非盐处理植株明显 ;Na Cl降低叶绿素含量 ,CO2 可使其轻微提高 ;盐对普通小麦TCAC中的异柠檬酸脱氢酶 (IDH)、琥珀酸脱氢酶 (SDH)、苹果酸脱氢酶 (MDH)和光呼吸中的乙醇酸氧化酶 (GO)、羟基丙酮酸还原酶 (HPR)有刺激作用 ,CO2 则抑制它们的活性。抗盐小麦对 CO2 /盐冲击的反应与普通小麦有差别。结果可以说明 ,CO2 能够减轻 Na Cl对植物的毒害效应     

Effect of fatty acids on growth of Japanese encephalitis virus cultivated in BHK-21 cells and phospholipid metabolism of the infected cells.

Growth of Japanese encephalitis virus (JEV) in BHK-21 cells was stimulated in the presence of 20 to 40 mug of the sodium salt of oleic acid (cis-9-octadecenoic acid, 9-18:1) per ml supplemented in Waymouth medium. The stimulatory effect of the salt was highest when 9-18:1 was added after adsorption of the virus. Study of the effect of other fatty acids on growth of JEV showed the following results: the longer the chain length of the saturated fatty acid salt, the higher the stimulatory effect on viral growth. In contrast, polyunsaturated fatty acids had an inhibitory effect on viral growth. The effect of isomeric cis-octadecenoic acids on viral growth was variable, depending upon the position of the double bond. The cis-6-octadecenoic acid had the highest inhibitory effect on growth of JEV compared to other isomeric octadecenoic acids. The sodium salt of (1-14C) cis-9-octadecenoic acid (9-18:1, 20 mug/ml) was rapidly incorporated into control and JEV-infected cells. Specific radioactivity in phosphatidylcholine dropped 12 to 24 h after virus inoculation, whereas synthesis of phosphatidylethanolamine increased 12 to 24 h after virus inoculation in infected cells compared to uninfected cells. Results from these studies suggest that phospholipid metabolism of infected cells is markedly changed, which can be associated with altered fatty acid metabolism when using labeled 9-18:1 fatty acid as a marker.     

Chemopreventive efficacy of selenium against N-nitrosodiethylamine-induced hepatoma in albino rats.

The chemopreventive/chemotherapeutic effect of sodium selenite on tricarboxylic acid cycle key enzymes was investigated against hepatoma induced by environmental carcinogen N-nitrosodiethylamine. Decreased activities of TCA cycle key enzymes such as isocitrate dehydrogenase (ICDH), succinate dehydrogenase (SDH), malate dehydrogenase (MDH) and alpha-ketoglutarate dehydrogenase (alpha-KGDH) in hepatoma and surrounding tissues of hepatoma-bearing rats were observed. Upon selenium supplementation the above biochemical changes were reverted in a dose- and duration-dependent manner. This study further confirms the chemopreventive/chemotherapeutic effect of sodium selenite which is found to be more effective in the initiation phase of carcinogenesis.     

Effects of salts on the halophilic alga Dunaliella viridis

Determinations of the salt sensitivity of enzymes extracted from the halophilic alga Dunaliella viridis revealed that pentose phosphate isomerase, ribulose diphosphate carboxylase, glucose-6-phosphate dehydrogenase, and phosphohexose isomerase were inhibited by NaCl concentrations far lower than that in the growth medium (3.75 m). The inhibition was reversible and was not prevented by preparing the extracts in the presence of salt. Potassium, lithium, and cesium chlorides were equally inhibitory. In contrast, whole cells require rather high levels of NaCl for optimal growth, whereas growth is inhibited by low levels of the other cations. The results suggest a specific mechanism for the exclusion of sodium from the interior of the cell.     

Influences of CO2/Salt Shock on Respiratory Enzyme Activities in Wheat Seedling with Different Salt Tolerance

Influences of CO2/salt shock on the growth status, chlorophyll contents and activities of enzymes including tricarboxylic acid cycle (isocitric dehydrogenase (IDH), malic dehydrogenase (MDH), succinic dehydrogenase (SDH)) and photorespiratory pathway (glycolate oxidase (GO), hydroxypyruvate reductase (HRR), catalase) in seedling of wheat with different salt tolerance have been studied. Results showed that the growth of two wheat varieties were inhibited by NaC1 and stimulated by CO2, the responses being greater in the salt-stressed than in the non-salt-stressed plants. The chlorophyll contents at two wheat varieties were decreased by NaC1 while chorophyll levels were elevated by CO2. The activities of SDH, IDH, MDH in TCAC, and GO, HPR in photorespiratory pathway were stimulated by NaCI in salt-sensitive wheat, whereas these enzymes activities were inhibited by CO2, their response being different in salt-tolerant wheat. Data suggested that increased CO2 levels might alleviate the adverse effects of salinity.     

Quantitative aspects of enzyme histochemistry on sections of freeze-substituted glycol methacrylate-embedded rat liver

Freeze-substituted rat liver embedded in glycol methacrylate (GMA) has been used to demonstrate the activities of several enzymes. The following enzymes could be detected in GMA-sections by the indicated histochemical procedure(s): 5-nucleotidase (lead salt, cerium-diaminobenzidine), alkaline phosphatase (indoxyl-tetrazolium salt), catalase (diaminobenzidine), acid phosphatase (diazonium salt), lactate dehydrogenase (tetrazolium salt) and glutamate dehydrogenase (tetrazolium salt). The activities of all these enzymes were dramatically decreased compared with the activities demonstrated in unfixed cryostat sections, with the exception of catalase. The activities of the following enzymes could not be detected in GMA-sections: glucose-6-phosphate dehydrogenase (tetrazolium salt), xanthine oxidoreductase (tetrazolium salt), d-amino acid oxidase (cerium-diaminobenzidine-cobalt-hydrogen peroxide) and glucose-6-phosphatase (cerium-diaminobenzidine). The possible role of restricted penetration of reagents into the resin was studied by measuring cytophotometrically the enzyme activities in GMA-sections of 3 and 6 m in thickness. For all the enzymes that could be detected, the 6 m : 3 m ratio varied from 1.4 to 2.7. An eventual retarded penetration of reagents into the resin was investigated by measuring cytophotometrically the amount of final reaction product during incubation for acid phosphatase and glutamate dehydrogenase activities. In both cases linear relationships without a lag phase were found for the specific enzyme activities with incubation time. Chemical denaturation of proteins or masking of active sites in proteins due to embedding in the resin monomer may be considered to be the main cause of decreased enzyme activities.     

The effects of storage on the retention of enzyme activity in cryostat sections. A quantitative histochemical study on rat liver

Summary The effect of storage of unfixed cryostat sections from rat liver for 4 h, 24 h, 3 days and 7 days at -25°C was studied on the activities of lactate dehydrogenase, glucose-6-phosphate dehydrogenase, xanthine oxidoreductase, glutamate dehydrogenase, succinate dehydrogenase (all demonstrated with tetrazolium salt procedures), glucose-6-phosphatase (cerium-diaminobenzidine method), 5-nucleotidase (lead salt method), dipeptidyl peptidase II, acid phosphatase (both simultaneous azo coupling methods), d-amino acid oxidase (cerium-diaminobenzidine-cobalt-hydrogen peroxide procedure) and catalase (diaminobenzidine method). The effect of drying of the cryostat sections at room temperature for 5 and 60 min was investigated as well. The enzyme activities were quantified by cytophotometric measurements of test and control reactions. The test minus control reaction was taken as a measure for specific enzyme activity. It was found that the activities of all the enzymes investigated, with one exception, were affected neither by storage of the cryostat sections at -25°C for up to 7 days, nor by drying of the sections at room temperature for up to 60 min. The exception was xanthine oxidoreductase, whose activity was reduced by 20% after 5 min drying of sections or after 4 h storage. Therefore, only incubations for xanthine oxidoreductase activity have to be performed immediately after cutting cryostat sections, whereas for the other enzymes a considerable margin appears to exist.     

Kinetic analysis of the inhibition of sulfate transport in human red blood cells by isothiocyanates

Summary A kinetic analysis of anion self-exchange in human red blood cells, in the presence of an irreversible inhibitor, is presented and applied to the study of the inactivation of sulfate transport by three isothiocyanates: 3-isothiocyano-1,5-naphthalenedisulfonic acid, disodium salt (INDS), 1-isothiocyano-4-naphthalene sulfonic acid, sodium salt, monohydrate (INS), and 1-isothiocyano-4-benzenesulfonic acid, sodium salt, monohydrate (IBS). The time dependence of the inhibition of sulfate transport by the isothiocyanates used could be described by a single exponential and could be shown to contain a reversible and an irreversible component. In each case a portion of sulfate efflux was found to be resistant to inactivation. The residual portion of the sulfate efflux varied with inhibition: 4% for INS, 16% for INDS, and 34% for IBS. INS showed the largest reversible inhibitory effect (12% of the flux remaining at 0.2mm inhibitor concentration), while INDS showed the weakest effect (92% of the flux remaining at 0.3mm inhibitor concentration). IBS had the highest rate of inactivation while INDS had the lowest. The kinetic analysis further suggests that all three isothiocyanates bind reversibly to an inhibitory site on the membrane before they bind covalently, and therefore irreversibly, to the same site on the membrane. The equilibrium constant for the dissociation of the reversibly-bound complex,K _i, and the rate of irreversible inactivation after all membrane sites are reversibly bound,k _max, have been computed for all three inhibitors: INDS (K _i=420m,k _max=5.04 hr^–1), INS (K _i=148 m,k _max=6.48 hr^–1), and IBS (K _i=208 m,k _max=8.11 hr^–1).     

Eco-physiological studies on Indian arid zone plants

Summary The separate and combined effects of sodium chloride and gibberellin (GA) on growth and the activities of alanine aminotransferase (GPT), aspartate aminotransferase (GOT) and glutamate dehydrogenase (GLDH) have been studied in the aerial parts of Pennisetum typhoides seedlings. Salt concentrations higher than 8.55×10³ M inhibited growth and reduced GLDH activity, but strongly stimulated the activity of GPT and, to a lesser extent, that of GOT. GA alone, on the other hand, stimulated growth but did not affect activity of any of the enzymes tested. In combination with salt, however, GA tended to counteract the effect of salt on both growth and enzyme activity. The possible significance of the results in explaining adaptation of plants under conditions of stress has been discussed.