Promotion of photosynthesis in transgenic rice over-expressing of maize C4 phosphoenolpyruvate carboxylase gene by nitric oxide donors

We determined the effects of exogenous nitric oxide on photosynthesis and gene expression in transgenic rice plants (PC) over-expressing the maize C₄ pepc gene, which encodes phosphoenolpyruvate carboxylase (PEPC). Seedlings were subjected to treatments with NO donors, an NO scavenger, phospholipase inhibitors, a Ca²⁺ chelator, a Ca²⁺ channel inhibitor, and a hydrogen peroxide (H₂O₂) inhibitor, individually and in various combinations. The NO donors significantly increased the net photosynthetic rate (P_N) of PC and wild-type (WT), especially that of PC. Treatment with an NO scavenger did inhibit the P_N of rice plants. The treatments with phospholipase inhibitors and a Ca²⁺ chelator decreased the P_N of WT and PC, and photosynthesis was more strongly inhibited in WT than in PC. Further analyses showed that the NO donors increased endogenous levels of NO and PLD activity, but decreased endogenous levels of Ca²⁺ both WT and PC. However, there was a greater increase in NO in WT and a greater increase in PLD activity and Ca²⁺ level in PC. The NO donors also increased both PEPC activity and pepc gene expression in PC. PEPC activity can be increased by SNP alone. But the expression of its encoding gene in PC might be regulated by SNP, together with PA and Ca²⁺.     

Docosahexaenoic acid enhances segregation of lipids between : 2H-NMR study

Solid-state ²H-NMR of [²H₃₁]-N-palmitoylsphingomyelin ([²H₃₁]16:0SM, PSM*), supplemented by differential scanning calorimetry, was used for the first time, to our knowledge, to investigate the molecular organization of the sphingolipid in 1:1:1 mol mixtures with 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (16:0-18:1PE, POPE) or 1-palmitoyl-2-docosahexaenoyl-sn-glycero-3-phosphoethanolamine (16:0-22:6PE, PDPE) and cholesterol. When compared with ²H-NMR data for analogous mixtures of [²H₃₁]16:0-18:1PE (POPE*) or [²H₃₁]16:0-22:6PE (PDPE*) with egg SM and cholesterol, molecular interactions of oleic acid (OA) versus docosahexaenoic acid (DHA) are distinguished, and details of membrane architecture emerge. SM-rich, characterized by higher-order, and PE-rich, characterized by lower-order, domains <20 nm in size are formed in the absence and presence of cholesterol in both OA- and DHA-containing membranes. Although acyl chain order within both domains increases on the addition of sterol to the two systems, the resultant differential in order between SM- and PE-rich domains is almost a factor of 3 greater with DHA than with OA. Our interpretation is that the aversion that cholesterol has for DHA—but not for OA—excludes the sterol from DHA-containing, PE-rich (nonraft) domains and excludes DHA from SM-rich/cholesterol-rich (raft) domains. We attribute, in part, the diverse health benefits associated with dietary consumption of DHA to an alteration in membrane domains.     

Partitioning of Triton X-100, deoxycholate and C10EO8 into bilayers composed of native and hydrogenated egg yolk sphingomyelin

We have used isothermal titration calorimetry (ITC) to study the thermodynamics of Triton X-100 (T_X-100), deoxycholate and decyl octaethylene glycol (C₁₀EO₈) penetration into bilayers composed of native (ESM) and hydrogenated egg yolk sphingomyelin (DHSM). Light scattering measurements were used to study the point of saturation (R_e,sat) and the onset of solubilization of membranes by the detergents. We found that DHSM bilayers at 25 °C were much more resistant to detergent partitioning (lower K) and gave higher reaction enthalpies (ΔH) for all three detergents compared to the ESM bilayer system. Because DHSM lacks double bonds (Δ^4trans and some cis bonds as well), attractive acyl chain interactions are favored in membranes of this lipid class. The high stability and cohesion of DHSM in membranes could be a crucial functional property of this lipid as it is enriched in eye lens membranes.     

Novel ruthenium(II) diamine complexes

The complex cations [Ru(C₇H₁₆N₂)(C₁₀H₁₄)Cl]⁺, [Ru(C₇H₁₆N₂)(C₆H₆)Cl]⁺, [Ru(C₉H₁₈N₂)(C₆H₆)Cl]⁺, [Ru(C₉H₁₈N₂)(C₁₀H₁₄)Cl]⁺ and [Ru(C₁₄H₁₆N₂)(C₁₀H₁₄)Cl]⁺ have been synthesised from the reaction between the ruthenium-arene complexes [with C₆H₆ (benzene) or C₁₀H₁₄ (p-cymene)] and the respective chiral diamines [C₇H₁₆N₂=(S)-(−)-2-aminomethyl-1-ethylpyrrolidine, C₉H₁₈N₂=(S)-(+)-2-(pyrrolidinylmethyl)-pyrrolidine, or C₁₄H₁₆N₂=(1R,2R)-(+)-1,2-diphenylethylenediamine], isolated and characterised as chloride salts using single-crystal X-ray diffraction. All complexes were fully characterised by elemental analysis, mass spectrometry, ¹³C and ¹H NMR, and also found to exhibit catalytic activity in the transfer hydrogenation of acetophenone to 1-phenylethanol at 50 °C (enantiomeric excesses range from ca. 25% to 60%, and conversions from ca. 30% to 50%).     

Differences in the pattern of attack of acidic,neutral, and basic phospholipases A2 of A. halys blomhofii on human erythrocyte membranes: Problems in interpretation of phospholipid location

Purified acidic (pI 4.9), neutral (pI 6.9), and basic (pI 8.7) phospholipase A₂ from Agkistrodon halys blomhofii showed characteristically different patterns of hemolysis and phospholipid hydrolysis of intact human erthyrocytes. Acidic and neutral enzymes were nonlytic in the early periods of incubations with intact erythrocytes whereas the basic enzyme caused immediate hemolysis (5–8%). Under nonlytic conditions acidic and neutral enzymes hydrolyzed only phosphatidyl choline (PC) (20 and 50%, respectively), whereas basic enzyme hydrolyzed not only PC (60%) but nearly 15% of the phosphatidylethanolamine (PE). Both PC and PE were hydrolyzed significantly when the three phospholipases A₂ were incubated individually with erythrocyte lysate or hypotonic ghosts (sealed or unsealed). The order of substrate preference for acidic and neutral enzymes was always PC > PE. On the contrary basic enzyme exhibited the property of substrate specificity reversal. It hydrolyzed PC faster than PE when the membranes were sealed whereas PE hydrolysis was faster than PC hydrolysis in unsealed membranes. Interestingly only the basic enzyme showed activity in the absence of Ca²⁺ and in the presence of 0.5 mm EDTA. Phospholipase C (Bacillus cereus or Clostridium perfringens) did not show the property of substrate specificity reversal although their ability to hydrolyze PC and PE was different. In general this study demonstrates the unique activity patterns of three physically different pure phospholipases A₂ on human erythrocyte membranes which could be of value in selectively modifying membrane phospholipids. In addition it also throws an important light on the fact that results obtained with phospholipases should be interpreted with caution particularly as regards the localization of phospholipids in membranes.     

CTP:phosphocholine cytidylyltransferase and protein kinase C recognize different physical features of membranes: differential responses to an oxidized phosphatidylcholine

Protein kinase C (PKC) and CTP:phosphocholine cytidylyltransferase (CT) are two examples of enzymes that are regulated by reversible binding to membranes, and this binding is influenced by membrane physical properties. CT activation by oxidized phosphatidylcholines was recently demonstrated and was linked to the acyl chain disordering effect of the oxidized species (Biochemistry 38, 15606). In this paper, we compare the responses of PKC and CT to an oxidized PC, and investigate the physical properties of lipid bilayers that modulate the activity of these enzymes. We show that 1-palmitoyl, 2-(11,15 dihydroxy) eicosatrienoyl PC (diOH-PAPC) caused less of an increase in the temperature of the lamellar to hexagonal II transition (T_H) of an unsaturated PE, compared to its parent, PAPC. Using a polarity-sensitive interfacial probe, we also found evidence to suggest that this oxidized PC increases interfacial packing pressure. We found that whereas diOH-PAPC activates CT, it inhibits PKC relative to the parent PAPC. The activities of both CT and PKC are known to increase in the presence of non-lamellar forming lipids. The greater activating effect of diOH-PAPC compared with PAPC, is consistent with a stimulation of the activity of CT by negative curvature strain. However, this is not the case with PKC, for which we suggest that surface packing pressure is of prime importance.     

The modulating effects of lipids on purified rat liver Golgi galactosyltransferase

Galactosyltransferase was purified from rat liver Golgi membranes. The Triton X-100, used to solubilize the enzyme was removed immediately prior to the lipid interaction studies. In lipid vesicles, prepared from a variety of phosphatidylcholines (PCs), including egg PC, DOPC, DMPC, DPPC and DSPC, the ability of the lipids to stimulate the enzyme decreased in the order egg PC greater than DOPC greater than DMPC greater than DPPC greater than DSPC, i.e. the lower the transition temperature (Tc) the greater the stimulation of the enzyme. A second, neutral lipid, phosphatidylethanolamine was used to permit a comparison of the effect of a different head group of the same net charge at neutral pH. The PEs included, egg PE, soy PE, Pl-PE, PE(PC) and DPPE in order of increasing Tc. The effect of the PEs was opposite to that of the PCs, i.e. the higher the Tc, the greater the stimulation of the enzyme. In fact egg PE and soy PE which have the lowest Tc values were inhibitory. Thus the modulation of the Golgi membrane galactosyltransferase by these lipids was different from that reported earlier for the bovine milk galactosyltransferase. The effects of two acidic lipids, egg phosphatidic acid (PA) and egg phosphatidylglycerol (PG) were studied also. Both totally inhibited the enzyme even at low concentrations of lipid, however, the PA was more effective than PG. In mixtures of neutral lipid (PC) and acidic lipid (PA or PG), the effect of the acidic lipid dominated. Even in the presence of excess PC, total inhibition of the enzyme was observed. It was concluded that the enzyme bound the acidic lipid preferentially to itself. The choice of the lipids allowed us to make several direct comparisons concerning the effect of the nature of the lipid head group on the activity of the enzyme. For example PE(PC), egg PA and egg PG would have fatty acid chains identical to egg PC since these three lipids are all prepared by modification of egg PC. As well, DPPE differs from DPPC only by nature of the head group. These comparisons indicated that not only the net charge but also chemical nature of the head group were important in the lipid modulation of Golgi galactosyltransferase.     

Thyroid Hormone Treatments Differentially Affect the Temperature Kinetics Properties of FoF1 ATPase and Succinate Oxidase as well as the Lipid/Phospholipid Profiles of Rat Kidney Mitochondria: A Correlative Study

Effect of thyroidectomy (Tx) and subsequent treatment with 3,5,3′-triiodo-l-thyronine (T₃) or replacement therapy (T_R) with T₃ + l-thyroxine (T₄) on the temperature kinetics properties of FoF₁ adenosine triphosphatase (ATPase, ATP synthase, H⁺-translocating ATP synthase EC 3.6.3.14) and succinate oxidase (SO) and on the lipid/phospholipid makeup of rat kidney mitochondria were examined. Tx lowered ATPase activity, which T₃ treatment restored. SO activity was unchanged in Tx but decreased further by T₃ treatment. T_R restored both activities. The energies of ATPase activation in the high and low temperature ranges (E _H and E _L) increased in the Tx and T₃ animals with decrease in phase transition temperature (Tt). T_R restored E _H and E _L but not Tt to euthyroid levels. E _H and E _L of SO decreased in Tx animals. T₃ and T_R restored E _H whereas E _L was restored only in the T_R group; Tt increased in both groups. Total phospholipid and cholesterol contents decreased significantly in Tx and T₃-treated animals. In Tx animals, sphingomyelin (SPM) and phosphatidylcholine (PC) components decreased, while phosphatidylserine (PS) and diphosphatidylglycerol components increased. T₃ and T_R treatments caused decreases in SPM, phosphatidylinositol and PS. PC and phosphatidylethanolamine (PE) increased in the T₃ group. T_R resulted in increased lysophospolipids and PE. Changes in kinetic parameters of the two enzymes were differently correlated with specific phospholipid components. Both T₃ and T_R regimens were unable to restore normal membrane structure-function relationships.     

The effect of plant hormone abscisic acid on model membranes - differential scanning calorimetry and planar bilayer membranes investigations

The effect of abscisic acid on the thermotropic properties of dipalmitoylphosphatidylcholine (DPPC) and on phosphatidylethanolamines (natural (PE) and dipalmitoylphosphatidylethanolamine (DPPE)) bilayers was investigated by differential scanning calorimetry (DSC). Abscisic acid eliminates the pretransition of DPPC, causes a downward shift of its temperature of melting (T_m) and broadens the melting peak without changing the enthalpy of melting. In natural PE bilayers interacting with abscisic acid a small decrease in the enthalpy of melting almost without change of T_m was detected, whereas in synthetic DPPE abscisic acid caused a small shift of T_m and small broadening of the melting peak without changing the enthalpy of melting. Abscisic acid increases the conductance to Na⁺ or K⁺ by three orders of magnitude in planar lipid membranes formed from PE monolayers and by less than two orders of magnitude in membranes formed from PC monolayers.     

Preservation of dried liposomes in the presence of sugar and phosphate

It has been well established that sugars can be used to stabilize liposomes during drying by a mechanism that involves the formation of a glassy state by the sugars as well as by a direct interaction between the sugar and the phospholipid head groups. We have investigated the protective effect of phosphate on solute retention and storage stability of egg phosphatidylcholine (egg PC) liposomes that were dried (air-dried and freeze-dried) in the presence of sugars and phosphate. The protective effect of phosphate was tested using both glucose (low T_g) and sucrose (high T_g) by measuring leakage of carboxyfluorescein (CF), which was incorporated inside the vesicles. Liposomes that were dried with glucose or phosphate alone showed complete leakage after rehydration. However, approximately 30% CF-retention was obtained using mixtures of phosphate and glucose. Approximately 75% CF-retention was observed with liposomes that were dried with sucrose. The solute retention further increased to 85% using mixtures of phosphate and sucrose. The pH of the phosphate buffer prior to drying was found to have a strong effect on the solute retention. Fourier transform infrared spectroscopy studies showed that phosphate and sugars form a strong hydrogen bonding network, which dramatically increased the T_g. The HPO₄²⁻ form of phosphate was found to interact stronger with sugars than the H₂PO₄⁻ form. The increased solute retention of liposomes dried in the sugar phosphate mixtures did not coincide with improved storage stability. At temperatures below 60 °C the rate of solute-leakage was found to be strikingly higher in the presence of phosphate, indicating that phosphate impairs storage stability of dried liposomes.     

Mobilization of arachidonic acid from phosphatidylethanolamine fraction to phosphatidylcholine fraction in platelets

Rabbit platelets rapidly incorporated methyl groups of [³H] methionine to phosphatidylcholine (PC). Rabbit platelets also incorporated [³H]choline to PC, but the rate of incorporation was far lower than that of [³H]methionine. Further fractionation of labeled PC revealed that a considerable amount of arachidonyl PC was synthesized via the N-methylation pathway. Thrombin stimulation resulted in a release of arachidonic acid from PC, and not from phosphatidylethanolamine (PE). These observations suggest that the N-methylation pathway plays an important role in the intracellular mobilization of arachidonic acid from the PE fraction to the PC fraction, this fraction being more sensitive to the hydrolysis with phospholipase A₂ during platelet activation.     

Asymmetric binuclear titanocenes linked by alkyl benzyl ethers: Synthesis, characterization and use as catalysts for ethylene polymerization

A series of novel asymmetric binuclear titanocenes linked with alkyl benzyl ethers p-[(C₅H₅TiCl₂)C₅H₄CH₂]C₆H₄O(CH₂)_n[C₅H₄(TiCl₂C₅H₅)] (n = 2-5) (13-16) have been synthesized by treating p-(LiC₅H₄CH₂)C₆H₄O(CH₂)_n(C₅H₄Li) (n = 2-5) (9-12) with C₅H₅TiCl₃. The new complexes have been characterized by elemental analysis and NMR spectra. Their catalytic activity for ethylene polymerization was investigated in the presence of aluminoxane (MAO). The results show that 13-16 are efficient catalysts for producing polyethylene (PE) with a broad molecular weight distribution (MWD). Their catalytic activity is highly dependent on the length of the alkyl chain and the polymerization conditions. A longer alkyl chain increases the catalytic activity, whereas the molecular weight of the produced polyethylene decreases.     

Fatty acid exchange of phospholipids in membranes of rat heart

Incorporation of ¹⁴C fatty acids in phospholipids of plasma membranes and sarcoplasmic reticulum of rat heart was studied. Mainly phosphatidylcholine (PC) and phosphatidylethanolamine (PE) were labelled. Our studies showed that the incorporation of unsaturated fatty acids (oleic and linoleic acid) was higher than for saturated fatty acids (palmitic and stearic acid). The range of uptake was between 0.2 and 2.2 nmol·mg⁻¹ protein·h⁻¹ and 0.5–7.4 nmol·μgatom⁻¹ P₁·h⁻¹, respectively. Uptake of activity in individual phospholipids (measured after separation on TLC) was calculated as percentage of total activity. Incorporation in phosphatidylcholine was higher than in phosphatidylethanolamine. Phosphatidylcholine showed an increasing sequence for the following fatty acids: C_18:0 < C_16:0 < C_18:0 < C_18:2. However, phosphatidylethanolamine showed a decreasing sequence for the incorporation of the same fatty acids. Labelling of PC was always greater than for PE, except for stearic acid which was better incorporated into phosphatidylethanolamine. Uptake of the same fatty acid into phospholipids of sarcoplasmic reticulum was always higher than uptake into plasma membranes. As incorporation of fatty acids bound to albumin was studied in isolated membranes of rat heart, the addition of ATP and CoASH was an absolute requirement.     

Nitrogen-fixing bacillus species from Egyptian soils: Acetylene reduction and cultural conditions

Summary Among 390 isolates from Egytiian soils initially grown on Brown's N-free agar, 15 facultative Bacillus isolates were able to reduce acetylene in Stanier's N-poor broth under both aerobic and anaerobic (N₂ atmosphere) conditions. Some of these isolates were Gram-positive, with unswollen sporangia and thin-walled endospores. Other strains were with slightly or definitely bulged sporangia. Yeast extract (0.01%) was essential for growth stimulation and N₂[C₂H₂] fixation by these isolates. Replacing yeast extract with 20 g/ml (NH₄)₂SO₄ or biotin, thiamine and amino acids (singly or in combination) resulted in stimulation of growth and N₂[C₂H₂] fixation, though at lower rates than in yeast extract.One isolate was able to grow and reduce C₂H₂ in Stanier's N-free liquid medium. Nitrogenase [C₂H₂] activity of the anaerobically grown and incubated cultures was greater than aerobic cultures. Addition of 0.1% CaCO₃ to the culture media significantly increased and O₂ partially inhibited, N₂[C₂H₂] fixation by these Bacillus isolates.Studies of the characteristics and N₂[C₂H₂] fixing activities of these isolates indicate that at least some of them are new nitrogen-fixingBacillus species.     

Effect of the structure of lipids favoring disordered domain formation on the stability of cholesterol-containing ordered domains (lipid rafts): identification of multiple raft-stabilization mechanisms

Despite the importance of lipid rafts, commonly defined as liquid-ordered domains rich in cholesterol and in lipids with high gel-to-fluid melting temperatures (T_m), the rules for raft formation in membranes are not completely understood. Here, a fluorescence-quenching strategy was used to define how lipids with low T_m, which tend to form disordered fluid domains at physiological temperatures, can stabilize ordered domain formation by cholesterol and high-T_m lipids (either sphingomyelin or dipalmitoylphosphatidylcholine). In bilayers containing mixtures of low-T_m phosphatidylcholines, cholesterol, and high-T_m lipid, the thermal stability of ordered domains decreased with the acyl-chain structure of low-T_m lipids in the following order: diarachadonyl > diphytanoyl > 1-palmitoyl 2-docosahexenoyl = 1,2 dioleoyl = dimyristoleoyl = 1-palmitoyl, 2-oleoyl (PO). This shows that low-T_m lipids with two acyl chains having very poor tight-packing propensities can stabilize ordered domain formation by high-T_m lipids and cholesterol. The effect of headgroup structure was also studied. We found that even in the absence of high-T_m lipids, mixtures of cholesterol with PO phosphatidylethanolamine (POPE) and PO phosphatidylserine (POPS) or with brain PE and brain PS showed a (borderline) tendency to form ordered domains. Because these lipids are abundant in the inner (cytofacial) leaflet of mammalian membranes, this raises the possibility that PE and PS could participate in inner-leaflet raft formation or stabilization. In bilayers containing ternary mixtures of PO lipids, cholesterol, and high-T_m lipids, the thermal stability of ordered domains decreased with the polar headgroup structure of PO lipids in the order PE > PS > phosphatidylcholine (PC). Analogous experiments using diphytanoyl acyl chain lipids in place of PO acyl chain lipids showed that the stabilization of ordered lipid domains by acyl chain and headgroup structure was not additive. This implies that it is likely that there are two largely mutually exclusive mechanisms by which low-T_m lipids can stabilize ordered domain formation by high-T_m lipids and cholesterol: 1), by having structures resulting in immiscibility of low-T_m and high-T_m lipids, and 2), by having structures allowing them to pack tightly within ordered domains to a significant degree.     

Nonallergenic urushiol derivatives inhibit the oxidation of unilamellar vesicles and of rat plasma induced by various radical generators

Urushiols consist of an o-dihydroxybenzene (catechol) structure and an alkyl chain of 15 or 17 carbons in the 3-position of a benzene ring and are allergens found in the family Anacardiaceae. We synthesized various veratrole (1,2-dimethoxybenzene)-type and catechol-type urushiol derivatives that contained alkyl chains of various carbon atom lengths, including –H, –C₁H₃, –C₅H₁₁, –C₁₀H₂₁, –C₁₅H₃₁, and –C₂₀H₄₁, and investigated their contact hypersensitivities and antioxidative activities. 3-Decylcatechol and 3-pentadecylcatechol displayed contact hypersensitivity, but the other compounds did not induce an allergic reaction, when the ears of rats were sensitized by treatment with the compounds every day for 20 days. Catechol-type urushiol derivatives (CTUDs) exerted very high radical-scavenging activity on the 1,1-diphenyl-2-picrylhydrazyl radical and inhibited lipid peroxidation in a methyl linoleate solution induced by 2,2′-azobis(2,4-dimethylvaleronitrile) (AMVN). However, veratrole-type urushiol derivatives did not scavenge or inhibit lipid peroxidation. CTUDs also acted as effective inhibitors of lipid peroxidation of the egg yolk phosphatidylcholine large unilamellar vesicle (PC LUV) liposome system induced by various radical generators such as AMVN, 2,2′-azobis(2-amidino-propane) dihydrochloride, and copper ions, although their efficiencies differed slightly. In addition, CTUDs suppressed formation of cholesteryl ester hydroperoxides in rat blood plasma induced with copper ions. CTUDs containing more than five carbon atoms in the alkyl chain showed excellent lipophilicity in a n-octanol/water partition experiment. These compounds also exhibited high affinities to the liposome membrane using the ultrafiltration method of the PC LUV liposome system. Therefore, CTUDs seem to act as efficient antioxidative compounds against membranous lipid peroxidation owing to their localization in the phospholipid bilayer. These results suggest that nonallergenic CTUDs act as antioxidants to protect against oxidative damage of cellular and subcellular membranes.     

Acetylene Reduction (Nitrogen Fixation) by Pulp and Paper Mill Effluents and by Klebsiella Isolated from Effluents and Environmental Situations

High rates of acetylene (C₂H₂) reduction (nitrogenase activity) were observed in woodroom effluent from a neutral sulfite semi-chemical mill under aerobic (up to 644 nmol of C₂H₄ produced per ml per h) and under anaerobic (up to 135 nmol of C₂H₄ produced per ml per h) conditions. Pasteurized effluent developed C₂H₂ reduction activity when incubated under anaerobic but not under aerobic conditions. Activities were increased by addition of 0.5 to 3.0% glucose or xylose. Enrichment and enumeration studies showed that N₂-fixing Azotobacter and Klebsiella were abundant, and N₂-fixing Bacillus was present. Of 129 isolates of Klebsiella from pulp mills, lakes, rivers, and drainage and sewage systems, 32% possessed nitrogen-fixing ability.     

Substrate-specificities of acid and alkaline ceramidases in fibroblasts from patients with Farber disease and controls

The specific activity of acid ceramidase (N-acylsphingosine deacylase, EC 3.5.1.23) was measured at pH4.5 in normal fibroblasts and in fibroblasts from patients with Farber disease and obligate heterozygotes. Greater activity was found when the synthetically made ceramide substrates contained shorter-chain fatty acids or higher content of double bonds. Acid ceramidase activities towards N-lauroyl- (C_12:0), N-myristoyl- (C_14:0) and N-palmitoyl- (C_16:0) sphingosine (C_18:1) were respectively about 38, 26 and 6 times higher than the activity towards the N-stearoyl (C_18:0) substrate. The activity towards N-linolenoylsphingosine (C_18:3/C_18:1), N-linoleoylsphingosine (C_18:2/C_18:1) and N-oleoylsphingosine (C_18:1/C_18:1) were respectively about 5, 4 and 3 times higher than the activity towards N-stearoylsphingosine (C_18:0/C_18:1). The activity towards N-stearoyldihydrosphingosine (C_18:0/C_18:0) was about 40% of that towards N-stearoylsphingosine. Fibroblast alkaline ceramidase possessed significant activity only towards ceramides of unsaturated fatty acids, with a pH optimum of about 9.0. Deficiency of acid ceramidase activity in fibroblasts from patients with Farber disease and intermediate activities in obligate heterozygotes were demonstrated with all ceramides examined except for N-hexanoylsphingosine (C_6:0/C_18:1), whereas alkaline ceramidase activity was unaffected. Comparative kinetic studies of acid ceramidase activity with N-lauroylsphingosine and N-oleoylsphingosine demonstrated about 5 (2–12)-fold and 7 (4–17)-fold higher K_m values in fibroblasts from patients with Farber disease as compared with normal controls. N-Lauroylsphingosine, towards which acid ceramidase activity in control fibroblasts was about 10 times higher than that towards N-oleoylsphingosine, may serve as a better substrate for enzymic diagnosis of Farber disease as well as for further characterization of the catalytically defective acid ceramidase.     

Synthesis and characterization of carbamoyl methyl pyrazole(CMPz) compounds of palladium(II) chloride: The crystal and molecular structure of [PdCl2{C3H3N2CH2CONBu2}2]

Carbamoyl methyl pyrazole compound of palladium(II) chloride of the type [PdCl₂L₂] (where L =  C₅H₇N₂CH₂CON(C₄H₉)₂, C₅H₇N₂CH₂CON(ⁱC₄H₉)₂, C₃H₃N₂CH₂CON(C₄H₉)₂, or C₃H₃N₂CH₂CON(ⁱC₄H₉)₂) has been synthesized and characterized by IR and ¹H NMR spectroscopy. The structure of the compound [PdCl₂{C₃H₃N₂CH₂CONⁱBu₂}₂] has been determined by single crystal X-ray diffraction and shows that the ligands are bonded through the soft pyrazolyl nitrogen atom to the palladium(II) chloride in a trans disposition.     

Acetylene Metabolism and Stimulation of Denitrification in an Agricultural Soil

The effects of C₂H₂ metabolism on N₂O production were examined in soil slurries. Enrichment of C₂H₂ consumption activity occurred only in aerobic incubations. Rapid disappearance of subsequent C₂H₂ additions, stimulation of CO₂ production, and most-probable-number enumerations of C₂H₂ utilizers indicated enrichment of the population responsible. During C₂H₂ consumption in slurries incubated statically under air, maximal rates of N₂O evolution were 19 times higher than those in anaerobic incubations. After 20 days of enrichment with C₂H₂, the production of N₂O by slurries supplemented with C₂H₂ and nitrate was 10 times higher than that in the unenriched controls. A Nocardia- or Arthrobacter-like bacterium was isolated that grew on C₂H₂ but did not denitrify. The behavior of soil inoculated with this bacterium became similar to that of C₂H₂-enriched soil incubated aerobically. Ethanol, acetate, and acetaldehyde were identified in enrichment experiments, and denitrification in soil slurries was stimulated by addition of the supernatant from a pure culture grown on mineral medium with C₂H₂. These results indicate that denitrification can be stimulated by the actions of an aerobic, nondenitrifying C₂H₂-metabolizing population. Utilization of intermediate metabolites by denitrifiers and enhanced O₂ consumption are two possible mechanisms for this stimulation.