3,3-Dimethyl-1-butanol, a parakairomone component to Aleurodicus dispersus (Hemiptera: Aleyrodidae)

C-6-based green leaf volatiles (GLVs) are signal molecules to herbivorous insects and play an important role in plant–herbivore interactions. How isomerization of GLVs affects insect’s olfactory response has been rarely tested. In laboratory and field experiments, we examined the effect of hexanol isomers on olfactory orientation of the spiraling whitefly, Aleruodicus dispersus Russell, a highly polyphagous pest. In a Y-tube oflactometer, we found that (±)-2-hexanol, 3-methyl-3-pentanol and 3,3-dimethyl-1-butanol significantly attracted female A. dispersus. The trap captures of 3,3-dimethyl-1-butanol were significantly more than that of (±)-2-hexanol and 3-methyl-3-pentanol, and its optimum concentration was 1 μ1/ml. We suggest that the anthropogenic compound 3,3-dimethyl-1-butanol can be exploited as a parakairomone (synthetic analogues of kairomone) to monitor and control adult A. dispersus.     

Amino Acid Sequences of Glycopeptides from Cucumisin

Oxidation of short-chain iso-alkanes (isobutane, isopentane, 2-methylpentane, and 3-methylpentane) was studied with propane-grown resting mycelia of Scedosporium sp. A-4. Isobutane was oxidized to terf-butanol, but both isobutane and tert-butanol were not used for growth. Isopentane was oxidized to 3-methyl-1-butanol, 2-methyl-2-butanol, and 3-methyl-2-butanol but not to 2-methyl-1-butanol. 2-Methylpentane was oxidized to 4-methyl-1-pentanol, 2-methyl-2-pentanol, and 4-methyl-2-pentanol but not to 2-methyl-1-pentanol or 2-methyl-3-pentanol. 3-Methylpentane was not oxidized. Oxidation of branched alcohols was also studied.     

Comparison of substrate specificity of alcohol dehydrogenases from human liver, horse liver, and yeast towards saturated and 2-enoic alcohols and aldehydes

The saturated and 2-enoic primary alcohols and aldehydes, ethanol, 1-propanol, 1-butanol, 3-methyl-1-butanol, 1-hexanol, phenylmethanol, 3-phenyl-1-propanol, 2-propen-1-ol, 2-buten-1-ol, 3-methyl-2-buten-1-ol, 2-hexen-1-ol, 3-phenyl-2-propen-1-ol, ethanal, 1-propanal, 1-butanal, 1-hexanal, phenylmethanal, 3-phenyl-1-propanal, 2-propen-1-al, 2-buten-1-al, 2-hexen-1-al, and 3-phenyl-2-propen-1-al, have been compared under uniform conditions as substrates for the alcohol dehydrogenase enzymes from horse and human liver and from yeast. Kinetic constants (K_m arid V) have been measured for each of the substrates with each of the enzymes; equilibrium constants for the various alcohol-aldehyde pairs have also been estimated. The results obtained emphasize the similarities of yeast alcohol dehydrogenase to horse and human liver alcohol dehydrogenase, showing the specificity of yeast alcohol dehydrogenase to be less restricted than formerly believed. In general, the 2-enoic alcohols are better substrates for all three alcohol dehydrogenases than their saturated analogs; on the other hand, saturated aldehydes are better substrates than the 2-enoic aldehydes. Based on these various findings, it is suggested that a more likely candidate than ethanol for the physiological substrate of alcohol dehydrogenase in mammalian systems may well be an unsaturated alcohol, although the wide variety of substrates catalyzed at high rates is not incompatible with a general detoxifying function for alcohols or aldehydes, or both, by alcohol dehydrogenase.     

Synthesis of Short-Chain Diols and Unsaturated Alcohols from Secondary Alcohol Substrates by the Rieske Nonheme Mononuclear Iron Oxygenase MdpJ

The Rieske nonheme mononuclear iron oxygenase MdpJ of the fuel oxygenate-degrading bacterial strain Aquincola tertiaricarbonis L108 has been described to attack short-chain tertiary alcohols via hydroxylation and desaturation reactions. Here, we demonstrate that also short-chain secondary alcohols can be transformed by MdpJ. Wild-type cells of strain L108 converted 2-propanol and 2-butanol to 1,2-propanediol and 3-buten-2-ol, respectively, whereas an mdpJ knockout mutant did not show such activity. In addition, wild-type cells converted 3-methyl-2-butanol and 3-pentanol to the corresponding desaturation products 3-methyl-3-buten-2-ol and 1-penten-3-ol, respectively. The enzymatic hydroxylation of 2-propanol resulted in an enantiomeric excess of about 70% for the (R)-enantiomer, indicating that this reaction was favored. Likewise, desaturation of (R)-2-butanol to 3-buten-2-ol was about 2.3-fold faster than conversion of the (S)-enantiomer. The biotechnological potential of MdpJ for the synthesis of enantiopure short-chain alcohols and diols as building block chemicals is discussed.     

Production of Methyl Ketones from Secondary Alcohols by Cell Suspensions of C2 to C4n-Alkane-Grown Bacteria

Nineteen new C₂ to C₄n-alkane-grown cultures were isolated from lake water from Warinanco Park, Linden, N.J., and from lake and soil samples from Bayway Refinery, Linden, N.J. Fifteen known liquid alkane-utilizing cultures were also found to be able to grow on C₂ to C₄n-alkanes. Cell suspensions of these C₂ to C₄n-alkane-grown bacteria oxidized 2-alcohols (2-propanol, 2-butanol, 2-pentanol, and 2-hexanol) to their corresponding methyl ketones. The product methyl ketones accumulated extracellularly. Cells grown on 1-propanol or 2-propanol oxidized both primary and secondary alcohols. In addition, the activity for production of methyl ketones from secondary alcohols was found in cells grown on either alkanes, alcohols, or alkylamines, indicating that the enzyme(s) responsible for this reaction is constitutive. The optimum conditions for in vivo methyl ketone formation from secondary alcohols were compared among selected strains: Brevibacterium sp. strain CRL56, Nocardia paraffinica ATCC 21198, and Pseudomonas fluorescens NRRL B-1244. The rates for the oxidation of secondary alcohols were linear for the first 3 h of incubation. Among secondary alcohols, 2-propanol and 2-butanol were oxidized at the highest rate. A pH around 8.0 to 9.0 was found to be the optimum for acetone or 2-butanone formation from 2-alcohols. The temperature optimum for the production of acetone or 2-butanone from 2-propanol or 2-butanol was rather high at 60°C, indicating that the enzyme involved in the reaction is relatively thermally stable. Metal-chelating agents inhibit the production of methyl ketones, suggesting the involvement of a metal(s) in the oxidation of secondary alcohols. Secondary alcohol dehydrogenase activity was found in the cell-free soluble fraction; this activity requires a cofactor, specifically NAD. Propane monooxygenase activity was also found in the cell-free soluble fraction. It is a nonspecific enzyme catalyzing both terminal and subterminal oxidation of n-alkanes.     

Microbial Oxidation of Gaseous Hydrocarbons: Production of Secondary Alcohols from Corresponding n-Alkanes by Methane-Utilizing Bacteria

Over 20 new strains of methane-utilizing bacteria were isolated from lake water and soil samples. Cell suspensions of these and of other known strains of methane-utilizing bacteria oxidized n-alkanes (propane, butane, pentane, hexane) to their corresponding secondary alcohols (2-propanol, 2-butanol, 2-pentanol, 2-hexanol). The product secondary alcohols accumulated extracellularly. The rate of production of secondary alcohols varied with the organism used for oxidation. The average rate of 2-propanol, 2-butanol, 2-pentanol, and 2-hexanol production was 1.5, 1.0, 0.15, and 0.08 μmol/h per 5.0 mg of protein in cell suspensions, respectively. Secondary alcohols were slowly oxidized further to the corresponding methylketones. Primary alcohols and aldehydes were also detected in low amounts (rate of production were 0.05 to 0.08 μmol/h per 5.0 mg of protein in cell suspensions) as products of n-alkane (propane and butane) oxidation. However, primary alcohols and aldehydes were rapidly metabolized further by cell suspensions. Methanol-grown cells of methane-utilizing bacteria did not oxidize n-alkanes to their corresponding secondary alcohols, indicating that the enzymatic system required for oxidation of n-alkanes was induced only during growth on methane. The optimal conditions for in vivo secondary alcohol formation from n-alkanes were investigated in Methylosinus sp. (CRL-15). The rate of 2-propanol and 2-butanol production was linear for the 40-min incubation period and increased directly with cell protein concentration up to 12 mg/ml. The optimal temperature and pH for the production of 2-propanol and 2-butanol were 40°C and pH 7.0. Metalchelating agents inhibited the production of secondary alcohols. The activities for the hydroxylation of n-alkanes in various methylotrophic bacteria were localized in the cell-free particulate fractions precipitated by centrifugation between 10,000 and 40,000 × g. Both oxygen and reduced nicotinamide adenine dinucleotide were required for hydroxylation activity. The metal-chelating agents inhibited hydroxylation of n-alkanes by the particulate fraction, indicating the involvement of a metal-containing enzyme system in the oxidation of n-alkanes. The production of 2-propanol from the corresponding n-alkane by the particulate fraction was inhibited in the presence of methane, suggesting that the subterminal hydroxylation of n-alkanes may be catalyzed by methane monooxygenase.     

Pentynyl dextran as a support matrix for immobilization of serine protease subtilisin Carlsberg and its use for transesterification of N-acetyl-l-phenylalanine ethyl ester in organic media

In this study, serine protease (subtilisin Carlsberg) was immobilized on pentynyl dextran (PyD, O–alkynyl ether of dextran, 1) and used for the transesterification of N-acetyl-l-phenylalanine ethyl ester (2) with different aliphatic (1-propanol, 1-butanol, 1-pentanol, 1-hexanol) and aromatic (benzyl alcohol, 2-phenyl ethanol, 4-phenyl-1-butanol) alcohols in tetrahydrofuran (THF). The effect of carbon chain length in aliphatic and aromatic alcohols on initial and average transesterification rate, transesterification activity of immobilized enzyme and yield of the reaction under selected reaction conditions was investigated. The transesterification reactivity of the enzyme and yield of the reaction increased as the chain length of the alcohols decreased. Furthermore, almost no change in yield was observed when the immobilized enzyme was repeatedly used for selected alcohols over six cycles. Intrinsic fluorescence analysis showed that the catalytic activity of the immobilized enzyme in THF was maintained due to retention of the tertiary structure of the enzyme after immobilization on PyD (1).     

Stearoyl paratoluenesulfonate. A powerful acylating agent for lipid synthesis

The utility of the mixed carboxylic-sulfonic acid anhydride stearoyl p-toluenesulfonate as a powerful, mild acylating agent for lipid synthesis is shown by the synthesis of rac 1,2-distearoyl-3-iodopropane, lecithin and a spin-labeled choline derivative from the corresponding alcohols. The method constitutes a significant improvement of earlier acylating methods.     

Studies on Flavor Components in Soybean

Volatile neutral fraction of flavor components in raw soybean was investigated. Ground raw soybean was packed in a glass column, warmed at about 50°C by a water-jacket column, and swept with nitrogen gas. Volatile materials swept out were trapped in bottles dipped in ice-water, solid carbon dioxide-acetone and liquid nitrogen in this order, and were submitted to gas chromatography. The peaks corresponding to basic and carbonyl compounds were eliminated by treating the original volatile materials with 2, 4-dinitro-phenylhydrazine in phosphoric acid, and the residual peaks were compared with authentic alcohols and esters.

Methanol, ethanol, 2-pentanol, isopentanol, n-pentanol, n-hexanol and n-heptanol were found as alcohols, and n-pentanol acetate was found as an ester. Other methods of identification of alcohols were carried out by introducing these into 3, 5-dinitrobenzoate derivatives, which were investigated by thin-layer and paper chromatographies.

Isopentanol, n-hexanol and n-heptanol were considered to be important factors among the volatile neutral components, which gave green bean-like odor to soybean.     

Cholesterol autoxidation: identification of the volatile fragments.

The volatile fragments of air-aged cholesterol were analysed by means of gas chromatography-mass spectrometry; The following fourteen compounds were identified: ethanol, acetic acid, acetone, 2-methylpropene, 2-methyl-1-propanol, 2-methyl-2-propanol, 2-butanone, 2-methylpropionic acid, 2-methyl-2-butanol, 2-pentanone, 3-methyl-2-butanone, 2-methyl-1-pentene, 2-methyl-2-pentanol, and 2-methyl-4-penten-2-ol. Their formation via decomposition of initially formed sterol hydroperoxides is discussed.     

Substrate entropy in enzyme enantioselectivity: an experimental and molecular modeling study of a lipase

The temperature dependence of the enantioselectivity of Candida antarctica lipase B for 3-hexanol, 2-butanol, 3-methyl-2-butanol, 3,3-dimethyl-2-butanol, and 1-bromo-2-butanol revealed that the differential activation entropy, deltaR-SdeltaS, was as significant as the differential activation enthalpy, DeltaR-SdeltaH, to the enantiomeric ratio, E. 1-Bromo-2-butanol, with isosteric substituents, displayed the largest deltaR-SdeltaS. 3-Hexanol displayed, contrary to other sec-alcohols, a positive deltaR-SdeltaS. In other words, for 3-hexanol the preferred R-enantiomer is not only favored by enthalpy but also by entropy. Molecular dynamics (MD) simulations and systematic search calculations of the substrate accessible volume within the active site revealed that the (R)-3-hexanol transition state (TS) accessed a larger volume within the active site than the (S)-3-hexanol TS. This correlates well with the higher TS entropy of (R)-3-hexanol. In addition, this enantiomer did also yield a higher number of allowed conformations, N, from the systematic search routines, than did the S-enantiomer. The substrate accessible volume was greater for the enantiomer preferred by entropy also for 2-butanol. For 3,3-dimethyl-2-butanol, however, neither MD-simulations nor systematic search calculations yielded substrate accessible volumes that correlate to TS entropy. Ambiguous results were achieved for 3-methyl-2-butanol.     

Volatile Compounds Originating from Mixed Microbial Cultures on Building Materials under Various Humidity Conditions

We examined growth of mixed microbial cultures (13 fungal species and one actinomycete species) and production of volatile compounds (VOCs) in typical building materials in outside walls, separating walls, and bathroom floors at various relative humidities (RHs) of air. Air samples from incubation chambers were adsorbed on Tenax TA and dinitrophenylhydrazine cartridges and were analyzed by thermal desorption-gas chromatography and high-performance liquid chromatography, respectively. Metabolic activity was measured by determining CO₂ production, and microbial concentrations were determined by a dilution plate method. At 80 to 82% RH, CO₂ production did not indicate that microbial activity occurred, and only 10% of the spores germinated, while slight increases in the concentrations of some VOCs were detected. All of the parameters showed that microbial activity occurred at 90 to 99% RH. The microbiological analyses revealed weak microbial growth even under drying conditions (32 to 33% RH). The main VOCs produced on the building materials studied were 3-methyl-1-butanol, 1-pentanol, 1-hexanol, and 1-octen-3-ol. In some cases fungal growth decreased aldehyde emissions. We found that various VOCs accompany microbial activity but that no single VOC is a reliable indicator of biocontamination in building materials.     

Effects of Temperature on Stereochemistry of Enzymatic Reactions

A brief discussion of the theoretical basis for effects of temperature on stereoselectivity of enzyme catalysed reactions is presented. In theory, the stereoselectivity of an enzymatic reaction can either increase or decrease as the reaction temperature is raised. The secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus reduces 2-butanone to (R)-2-butanol at 37° C, with increased stereoselectivity at higher temperatures and in the presence of NADP analogues. In contrast, at 37°, 2-pentanone and 2-hexanone are reduced to (S)-2-pentanol and (S)-2-hexanol, respectively, but the stereoselectivity decreases at higher temperatures and in the presence of NADP analogues. Reduction of racemic 2-methylbutanal by the primary alcohol dehydrogenase from T. ethanolicus gives (S)-2-methyl-1-butanol with greater stereospecificity at 35° (51% e.e.) than at 15° (14% e.e.). Horse liver alcohol dehydrogenase shows a preference for oxidation of the (S)-enantiomers of acyclic secondary alcohols at 25°, with a decrease in stereospecificity at higher temperatures.     

Olfactory discrimination of structurally similar alcohols by cockroaches

The capability of the cockroach Periplaneta americana to discriminate odors of structurally similar aliphatic alcohols was studied by using an operant conditioning paradigm. Cockroaches were trained to discriminate three odors: one odor associated with sucrose solution (reward) and two odors associated with NaCl solution (non-reward). After training, their odor preferences were tested by counting the number of visits to each odor source. We tested the capability of cockroaches to discriminate (1) three normal aliphatic alcohols with different numbers of carbon (1-pentanol, 1-hexanol and 1-octanol), (2) three C6 aliphatic alcohols (1-hexanol, 2-hexanol and trans-2-hexen-1-ol), (3) binary mixtures of two of these three alcohols and their components, and (4) 1-hexanol solution of three different concentrations (1, 10 and 100 micro g micro l(-1)). Cockroaches exhibited higher preferences for the odors associated with reward in these tests, and we therefore conclude that cockroaches can discriminate these odors. However, discrimination of 1-hexanol and trans-2-hexen-1-ol and their binary mixture was imperfect, in that some statistical tests suggested significant level of discrimination but other tests did not. In addition, the cockroaches learned to associate a 1-hexanol solution of the highest or lowest concentration with sucrose reward but failed to learn to associate 1-hexanol of an intermediate concentration with reward.     

Constituants volatils de Lycopersicon esculentum

The volatile components of the tomato have been studied by combined GC-MS with and without previous separation by preparative GC. 5-Methylfurfuryl ketone, furfuryl alcohol, p-anisaldehyde, p-vinylphenol and geraniol were identified and 2-methyl-2-pentanol, 2-heptanol, 4-heptanol, trans-3-hexene-1-ol and trans-2-hexene-1-ol, sec-butyl butyrate, lactone of 5,6-dihydroxyhexanoic acid and isopropyl anisole were tentatively identified.     

Rhizomorph Formation in Fungi

The effect on growth and rhizomorph formation of 12 alcohols (methanol, ethanol, 1-propanol, 2-propanol, 1-butanol, 2-butanol, iso-butyl alcohol, tert-butyl alcohol, 1-pentanol, iso-amyl alcohol, ethylene glycol and glycerol) at different concentrations has been examined for 2 isolates of Armillaria mellea (Vahl ex Fr.) Quél. and 1 of Clitocybe geotropa (Bull. ex Fr.) Quél. The fungi were cultivated for 28 days on a synthetic, liquid glucose medium with the alcohols as supplement. The following alcohols strongly stimulated growth and rhizomorph formation: ethanol, 1-propanol and 1-butanol. A great variation was demonstrated between the isolates in relation to rhizomorph production, morphology, and ability to be stimulated by different alcohols.     

Production of 2-methyl-1-butanol and 3-methyl-1-butanol in engineered Corynebacterium glutamicum

The pentanol isomers 2-methyl-1-butanol and 3-methyl-1-butanol represent commercially interesting alcohols due to their potential application as biofuels. For a sustainable microbial production of these compounds, Corynebacterium glutamicum was engineered for producing 2-methyl-1-butanol and 3-methyl-1-butanol via the Ehrlich pathway from 2-keto-3-methylvalerate and 2-ketoisocaproate, respectively. In addition to an already available 2-ketoisocaproate producer, a 2-keto-3-methylvalerate accumulating C. glutamicum strain was also constructed. For this purpose, we reduced the activity of the branched-chain amino acid transaminase in an available C. glutamicum l-isoleucine producer (K2P55) via a start codon exchange in the ilvE gene enabling accumulation of up to 3.67 g/l 2-keto-3-methylvalerate. Subsequently, nine strains expressing different gene combinations for three 2-keto acid decarboxylases and three alcohol dehydrogenases were constructed and characterized. The best strains accumulated 0.37 g/l 2-methyl-1-butanol and 2.76 g/l 3-methyl-1-butanol in defined medium within 48 h under oxygen deprivation conditions, making these strains ideal candidates for additional strain and process optimization.     

Role of the MexAB-OprM Efflux Pump of Pseudomonas aeruginosa in Tolerance to Tea Tree (Melaleuca alternifolia) Oil and Its Monoterpene Components Terpinen-4-ol, 1,8-Cineole, and α-Terpineol

Using a series of efflux mutants of Pseudomonas aeruginosa, the MexAB-OprM pump was identified as contributing to this organism's tolerance to the antimicrobial agent tea tree (Melaleuca alternifolia) oil and its monoterpene components terpinen-4-ol, 1,8-cineole, and α-terpineol. These data show that a multidrug efflux system of P. aeruginosa can extrude monoterpenes and related alcohols.     

Engineering of an Escherichia coli Strain for the Production of 3-Methyl-1-Butanol

3-Methyl-1-butanol is a potential fuel additive or substitute. Previously this compound was identified in small quantities in yeast fermentation as one of the fusel alcohols. In this work, we engineered an Escherichia coli strain to produce 3-methyl-1-butanol from glucose via the host's amino acid biosynthetic pathways. Strain improvement with the removal of feedback inhibition and competing pathways increased the selectivity and productivity of 3-methyl-1-butanol. This work demonstrates the feasibility of production of 3-methyl-1-butanol as a biofuel and shows promise in using E. coli as a host for production.     

Effects of Temperature on Stereochemistry of Enzymatic Reactions

A brief discussion of the theoretical basis for effects of temperature on stereoselectivity of enzyme catalysed reactions is presented. In theory, the stereoselectivity of an enzymatic reaction can either increase or decrease as the reaction temperature is raised. The secondary alcohol dehydrogenase from Thermoanaerobacter ethanolicus reduces 2-butanone to (R)-2-butanol at 37° C, with increased stereoselectivity at higher temperatures and in the presence of NADP analogues. In contrast, at 37°, 2-pentanone and 2-hexanone are reduced to (S)-2-pentanol and (S)-2-hexanol, respectively, but the stereoselectivity decreases at higher temperatures and in the presence of NADP analogues. Reduction of racemic 2-methylbutanal by the primary alcohol dehydrogenase from T. ethanolicus gives (S)-2-methyl-1-butanol with greater stereospecificity at 35° (51% e.e.) than at 15° (14% e.e.). Horse liver alcohol dehydrogenase shows a preference for oxidation of the (S)-enantiomers of acyclic secondary alcohols at 25°, with a decrease in stereospecificity at higher temperatures.