Stereochemical course of the generation of 3-mercaptohexanal and 3-mercaptohexanol by beta-lyase-catalyzed cleavage of cysteine conjugates

The product resulting from the reaction between E-2-hexenal and l-cysteine was shown to be a diastereoisomeric mixture of 2-(2-S-l-cysteinylpentyl)-1,3-thiazolidine-4-carboxylic acid 1. Treatment of the conjugate with two sources of cysteine-S-conjugate beta-lyase (tryptophanase from E. coli and a crude enzyme extract prepared from Eubacterium limosum) resulted in the formation of 3-mercaptohexanal. The reaction proceeded with a slight preference for the (S)-configured product, however, with low conversion rate. The role of 3-S-l-cysteinylhexanal 2 as substrate for beta-lyases was demonstrated by in situ generation of 2 from 3-S-(N-acetyl-l-cysteinyl)hexanal using acylase. Opposite enantioselectivity was observed for the liberation of 3-mercaptohexanol from 3-S-l-cysteinylhexanol 5 by the enzyme preparations from Eubacterium limosum and tryptophanase. Various yeasts produced 3-mercaptohexanol starting from 1 as well as from 5. The reactions proceeded without preferential formation of one of the enantiomers.     

Inhibition of enzymatic browning of chlorogenic acid by sulfur-containing compounds

The antibrowning activity of sodium hydrogen sulfite (NaHSO(3)) was compared to that of other sulfur-containing compounds. Inhibition of enzymatic browning was investigated using a model browning system consisting of mushroom tyrosinase and chlorogenic acid (5-CQA). Development of brown color (spectral analysis), oxygen consumption, and reaction product formation (RP-UHPLC-PDA-MS) were monitored in time. It was found that the compounds showing antibrowning activity either prevented browning by forming colorless addition products with o-quinones of 5-CQA (NaHSO(3), cysteine, and glutathione) or inhibiting the enzymatic activity of tyrosinase (NaHSO(3) and dithiothreitol). NaHSO(3) was different from the other sulfur-containing compounds investigated, because it showed a dual inhibitory effect on browning. Initial browning was prevented by trapping the o-quinones formed in colorless addition products (sulfochlorogenic acid), while at the same time, tyrosinase activity was inhibited in a time-dependent way, as shown by pre-incubation experiments of tyrosinase with NaHSO(3). Furthermore, it was demonstrated that sulfochlorogenic and cysteinylchlorogenic acids were not inhibitors of mushroom tyrosinase.     

Elucidation of the enantioselective enzymatic hydrolysis of chiral herbicide dichlorprop methyl by chemical modification

Up to 25% of the current pesticides are chiral, the molecules have chiral centers, but most of them are used as racemates. In most cases, enantiomers of chiral pesticides have different fates in the environment. Knowledge of the function of amino acids of enzymes involved in enantioselective behaviors contributes to the understanding of the enantioselectivity of chiral pesticides. In this work, Aspergillus niger lipase (ANL, EC3.1.1.3) was chemically modified using bromoacetic acid (BrAc), 2,3-butanedione (BD), N-bromosuccinimide (NBS), and methanal. The enantioselectivity of the enzymatic hydrolysis of 2,4-dichlorprop-methyl (DCPPM) was investigated by chiral GC. The results have suggested that histidine, arginine, and tryptophan are essential for lipase activity and might be involved in the catalytic site of ANL. In addition, histidine and lysine play an important role in determining the observed enantioselective hydrolysis of chiral herbicide dichlorprop methyl. The molecular modeling study revealed that the essential hydrogen bonds formed between DCPPM and catalytic residues of ANL might be responsible for the enantioselectivity of DCPPM. The loss of enantioselectivity can also arise from the fact that the modification of the amino acids may cause changes in both the nature of the ANL enzyme conformation and the binding pattern of DCPPM. Our study provides basic information for the exploration of the enantioselective interaction mechanism of enzymes with chiral pesticides.     

硅藻土吸附增强的(RS)-2,4-DP对映体选择性酶促水解

利用手性气相色谱技术研究了硅藻土吸附作用对(RS) 2 ,4 二氯苯氧丙酸甲酯(2 ,4 DP)酶促水解对映体选择性的影响。实验结果表明,硅藻土吸附作用显著增强了酶促反应的对映体选择性(ER值由1.5 8增加到5 .31)。进一步研究表明,硅藻土对脂肪酶的吸附,引起酶构象变化,影响农药底物与酶结合的微环境,是酶促反应的对映体选择性增强的主要原因。吸附在硅藻土上的脂肪酶,与R 2 ,4 DP结合更为困难,反应速率下降;而S 2 ,4 DP接近酶反应中心更加容易,反应速率上升。此外,硅藻土吸附作用对农药底物的束缚引起处于“自由状态”的底物减少,也使酶促反应的对映体选择性略有增强     

Smoke-derived taint in wine: the release of smoke-derived volatile phenols during fermentation of Merlot juice following grapevine exposure to smoke

The release of smoke-derived volatile phenols during the fermentation of Merlot grapes, following grapevine exposure to smoke, has been investigated. The concentrations of guaiacol, 4-methylguaiacol, 4-ethylguaiacol, 4-ethylphenol, and eugenol were determined by gas chromatography-mass spectrometry and found to increase throughout the winemaking process. Only trace levels (< or = 1 microg/L) of guaiacol and 4-methylguaiacol could be detected in free run juice derived from the fruit of smoked vines; the highest levels, 388 microg/L and 93 microg/L, respectively, were observed in the finished wine. Control wine (derived from fruit of unsmoked vines) contained 4 microg/L guaiacol, with the volatile phenols either not detected or detected at only trace levels (< or = 1 microg/L) throughout fermentation. The role of enzyme and acid catalyzed hydrolysis reactions in releasing smoke-derived volatile compounds was also investigated. The volatile phenols were released from smoked free run juice by strong acid hydrolysis (pH 1.0) and enzyme (beta-glucosidase) hydrolysis, but not mild acid hydrolysis (juice pH 3.2-3.7). Guaiacol was again the most abundant smoke-derived phenol, present at 431 microg/L and 325 microg/L in strong acid and enzyme hydrolysates, respectively. Only trace levels of each phenol could be detected in each control hydrolysate. This study demonstrates the potential for under-estimation of smoke taint in fruit and juice samples; the implications for the assessment of smoke taint and quantification of volatile phenols are discussed.     

Characterization of the most odor-active compounds of Iberian ham headspace

Gas chromatography-olfactometry (GC-O) based on detection frequency (DF) was used to characterize the most odor-active compounds from the headspace of Iberian ham. Twenty-eight odorants were identified by GC-O on two capillary columns, including aldehydes (11), sulfur-containing compounds (7), ketones (5), nitrogen-containing compounds (2), esters (2), and an alcohol. Among them, the highest odor potencies (DF values) were found for 2-methyl-3-furanthiol, 2-heptanone, 3-methylbutanal, methanethiol, hexanal, hydrogen sulfide, 1-penten-3-one, 2-methylpropanal, ethyl 2-methylbutyrate, and (E)-2-hexenal. Nine of the 28 most odor-active compounds were identified for the first time as aroma components of dry-cured ham, including hydrogen sulfide, 1-penten-3-one, (Z)-3-hexenal, 1-octen-3-one, and the meaty-smelling compounds 2-methyl-3-furanthiol, 2-furfurylthiol, 3-mercapto-2-pentanone, 2-acetyl-1-pyrroline, and 2-propionyl-1-pyrroline.     

Aroma extract dilution analysis of a beeflike process flavor from extruded enzyme-hydrolyzed soybean protein

Aroma-active compounds from a beeflike process flavor, produced by extrusion of enzyme-hydrolyzed vegetable protein (E-HVP), were analyzed using aroma extract dilution analysis. The number of aroma-active compounds and the aroma intensity were increased by the addition of aroma precursors prior to extrusion. The most intense compound was 2-methyl-3-furanthiol having a cooked rice/vitamin-like/meaty aroma note. Several sulfur-containing furans, such as 2-methyl-3-(methylthio)furan, 2-methyl-3-(methyldithio)furan, and bis(2-methylfuryl)disulfide, were detected with high flavor dilution (FD) factors. Some pyrazines, such as 2-ethyl-3,5-dimethylpyrazine, 2,6-diethylpyrazine, and 3,5-diethyl-2-methylpyrazine, also had high FD factors. It is hypothesized that sulfur-containing amino acids and thiamin were important precursors in aroma formation in process flavor from E-HVP.     

Amino-N compounds found in soil organic matter hydrolysates of a loamy sand using an immobilized protease reactor column

Summary Soil organic matter (OM) from seven different fertility plots of a loamy sand was extracted and fractionated into high- and low-molecular-weight (HMW, LMW) fractions using gel filtration. The fractions were acid-hydrolyzed to determine the amino sugar and amino acid contents. The same fractions were hydrolyzed with an immobilized protease reactor column. Reverse-phase high-performance liquid chromatography (HPLC) was used to identify the soil amino-N compounds. With the HMW fraction as substrate, the enzyme released less than 1% of 11 amino-N compounds determined by acid hydrolysis. Phenylalanine and leucine, however, were recovered in quantities of 2% and 4%, respectively. Immobilized protease hydrolysis of the LMW fraction recovered considerably more amino-N compounds compared with acid hydrolysis of the same fractions. Each system of hydrolysis produced some amino-N compounds not found in the other. We conclude that an immobilized enzyme reactor column will allow a researcher to perform time-course hydrolysis, so that hydrolysis intermediates, e.g. peptides, can be separated and identified.     

Purification and characterization of a phytate-degrading enzyme from germinated faba beans (Vicia faba Var. Alameda)

A phytate-degrading enzyme was purified approximately 2190-fold from germinated 4-day-old faba bean seedlings to apparent homogeneity with a recovery of 6% referred to the phytase activity in the crude extract. It behaves as a monomeric protein of a molecular mass of approximately 65 kDa. The phytate-degrading enzyme belongs to the acidic phytases. It exhibits a single pH optimum at 5.0. Optimal temperature for the degradation of sodium phytate is 50 degrees C. Kinetic parameters for the hydrolysis of sodium phytate are K(M) = 148 micromol L(-1) and k(cat) = 704 s(-1) at 35 degrees C and pH 5.0. The faba bean phytase exhibits a broad affinity for various phosphorylated compounds and hydrolyzes phytate in a stepwise manner. The first hydrolysis product was identified as D/L-myo-inositol(1,2,3,4,5)pentakisphosphate.     

Volatile components in crabmeats of Charybdis feriatus

The volatile components of different meats (legs with claws, body, and carapace) of a popularly consumed edible crab in Asia, Charybdis feriatus, were investigated. Samples were extracted by simultaneous steam distillation-solvent extraction and analyzed by gas chromatography/mass spectrometry. Among 177 compounds detected, 130 were positively identified. Seventy-six compounds were previously reported in other crab species. A greater number of naphthalenes were detected in this crab compared with other crabs in the literature. Aromatic compounds, alcohols, and sulfur-containing compounds were the three predominant groups with >15 components. Carapace tissue contained a greater number of volatile components in each group, except for sulfur-containing compounds. Most of the common components in the leg meat and the body meat were found at similar levels (p > 0.05). Carapace tissue generally had the highest quantity of common components among the meats. The higher levels of volatile components present in the carapace tissue might account for its stronger flavor compared with the other meats. Furthermore, the higher number of aldehydes and lower number of sulfur-containing compounds detected in the carapace meat might contribute to its unique flavor.     

Investigation of bound aroma constituents of yellow-fleshed nectarines (Prunus persica L. Cv. Springbright). changes in bound aroma profile during maturation

Glycosidically bound volatile constituents of yellow-fleshed clingstone nectarines (cv. Springbright) were identified and quantified at three stages of maturity. Glycoconjugates were isolated by LC on a C(18) reversed phase column with methanol elution followed by hydrolysis with a commercial pectinase enzyme. Forty-five bound aglycons were identified for the first time in yellow-fleshed nectarine. Thirty were terpene derivatives, and the most abundant ones were (E)- and (Z)-furan linalool oxides, linalool, alpha-terpineol, (E)-pyran linalool oxide, 3,7-dimethylocta-1,5-diene-3,7-diol, linalool hydrate, 8-hydroxy-6,7-dihydrolinalool, (E)- and (Z)-8-hydroxylinalools, and (E)- and (Z)-8-hydroxygeraniols. The group of C(13) norisoprenoids included 3-hydroxy-beta-damascone, 3-hydroxy-7,8-dihydro-beta-ionone, 3-oxo-alpha-ionol, 3-hydroxy-7,8-dihydro-beta-ionol, 3-hydroxy-beta-ionone, 3-oxo-7,8-dihydro-alpha-ionol, 3-hydroxy-5,6-epoxy-beta-ionone, 3-oxo-retro-alpha-ionol (isomers I and II), 3-hydroxy-7,8-dehydro-beta-ionol, 4,5-dihydrovomifoliol, and vomifoliol. Generally, levels of bound compounds, in particular monoterpenols and C(13) norisoprenoids, increased significantly with maturation. delta-Decalactone was the only lactone found in the enzymatic hydrolysate of yellow-fleshed nectarine, but its level was much lower than that of its free form.     

Enzymatic production of xylooligosaccharides from cotton stalks

Xylooligosaccharide (XO) production was performed from xylan, which was obtained by alkali extraction from cotton stalk, a major agricultural waste in Turkey. Enzymatic hydrolysis was selected to prevent byproduct formation such as xylose and furfural. Xylan was hydrolyzed using a commercial xylanase preparation, and the effects of pH, temperature, hydrolysis period, and substrate and enzyme concentrations on the XO yield and degree of polymerization (DP) were investigated. Cotton stalk contains about 21% xylan, the composition of which was determined as 84% xylose, 7% glucose, and 9% uronic acid after complete acid hydrolysis. XOs in the DP range of 2-7 (X6 approximately X5>X2>X3) were obtained with minor quantities of xylose in all of the hydrolysis conditions used. Although after 24 h of hydrolysis at 40 degrees C, the yield was about 53%, the XO production rate leveled off after 8-24 h of hydrolysis. XO yield was affected by all of the parameters investigated; however, none of them affected the DP of the end product significantly, except the hydrolysis period. Enzyme hydrolysis was maintained by the addition of fresh substrate after 72 h of hydrolysis, indicating the persistence of enzyme activity. The optimal hydrolysis conditions were determined as 40 degrees C, pH 5.4, and 2% xylan. The obtained product was fractionated via ultrafiltration by using 10, 3, and 1 kDa membranes. Complete removal of xylanase and unhydrolyzed xylan was achieved without losing any oligosaccharides having DP 5 or smaller by 10 kDa membrane. After a two-step membrane processing, a permeate containing mostly oligosaccharides was obtained.     

Stability of thiols in an aqueous process flavoring

The flavor stability of an aqueous solution of a savory model process flavoring based on ribose and cysteine was investigated during accelerated storage at 50 degrees C. Of the three sulfur-containing flavor-impact components investigated, 2-methyl-3-furanthiol was found to be the least stable (59% decrease/24 h), and it was followed by 2-furfurylthiol (28% decrease/24 h), 2-mercapto-3-butanone (14% decrease/24 h), and 2,5-dimethyl-4-hydroxy-3(2H)-furanone (max. 10% decrease/24 h). Both cysteine and ribose were found to affect the stability of various flavor compounds. A mechanism for the instability of 2-methyl-3-furanthiol is proposed, and was confirmed by H-D exchange experiments.     

Isolation and identification of a red pigment from Allium subgenus Melanocrommyum

Three new sulfur-containing compounds were identified in Allium L. species belonging to the subgenus Melanocrommyum as the first examples of sulfur-containing pyrrole derivatives in nature. Some of these species are traditionally used in Southwest and Central Asia as vegetables and herbal drugs. A hypothetical biogenetic scheme is proposed in which L-(+)- S-(3-pyrrolyl)cysteine sulfoxide is enzymically degraded. The resulting 2-lactyl-3'-pyrrolyl sulfoxide is condensed readily to the red pigment 3,3'-dithio-2,2'-dipyrrole. All compounds are chemically unstable, rendering the analysis extremely difficult. Correlation NMR in combination with diffusion NMR allowed the identification of these low molecular weight compounds. For the first time, the compounds involved in the coloring process of Allium plant material have been identified from native plant material.     

Preparation and characterization of papain-modified sesame (Sesamum indicum L.) protein isolates

Defatted sesame meal ( approximately 40-50% protein content) is very important as a protein source for human consumption due to the presence of sulfur-containing amino acids, mainly methionine. Sesame protein isolate (SPI) is produced from dehulled, defatted sesame meal and used as a starting material to produce protein hydrolysate by papain. Protein solubility at different pH values, emulsifying properties in terms of emulsion activity index (EAI) and emulsion stability index (ESI), foaming properties in terms of foam capacity (FC) and foam stability (FS), and molecular weight distribution of the SPI hydrolysates were investigated. Within 10 min of hydrolysis, the maximum cleavage of peptide bonds occurred as observed from the degree of hydrolysis. Protein hydrolysates have better functional properties than the original SPI. Significant increase in protein solubility, EAI, and ESI were observed. The greatest increase in solubility was observed between pH 5.0 and 7.0. The molecular weight of the hydrolysates was also reduced significantly during hydrolysis. These improved functional properties of different protein hydrolysates would make them useful products, especially in the food, pharmaceutical, and related industries.     

Volatiles from roasted byproducts of the poultry-processing industry

Volatiles of roasted chicken breast muscle and byproducts, such as backbones, breastbones, spent bones, and skin, were investigated. Total volatile concentrations ranged from 2030 ppb in the roasted backbones to 4049 ppb in the roasted skin. The major classes of volatile compounds detected in roasted samples were aldehydes (648-1532 ppb) and alcohols (336-1006 ppb). Nitrogen- and/or sulfur-containing compounds were also detected in appreciable quantities (161-706 ppb) in all samples. For all samples, hexanal and 2-methyl-2-buten-1-ol were dominant among the aldehydes and alcohols, respectively. Among the nitrogen- and sulfur-containing compounds, Maillard reaction products, such as tetrahydropyridazines, piperidines, and thiazoles, were the major contributors to the total volatile content in all samples. The composition of volatiles observed in roasted byproducts was markedly different from that of the roasted breast muscle. Therefore, the blending of the byproducts in appropriate proportions or blending of volatile flavor extracts from different byproducts may be necessary to obtain an aroma that mimics roasted chicken aroma.     

Quantitative analysis of cystine, methionine, lysine, and nine other amino acids by a single oxidation--4 hour hydrolysis method

The sulfur-containing amino acids cystine and methionine play important roles in animal, especially avian, nutrition. Because these sulfur-containing amino acids are destroyed to varying extents by 6N HCl hydrolysis, oxidation and hydrolysis of cystine to cysteic acid and methionine to methionine sulfone have been widely used for determination of cystine and methionine. Lysine is considered the next limiting amino acid after the sulfur amino acids in poultry nutrition; therefore, determination of the amino acid content of rations focuses first on these 3 amino acids. The objective of this investigation was to establish whether lysine and other amino acids could be accurately determined in proteinaceous materials which had undergone performic acid oxidation. To perform this evaluation, lysine was determined in a variety of protein-containing materials both with and without performic acid oxidation. Performic acid oxidation followed by 6N HCl hydrolysis at 145 degrees C for 4 h allows accurate measurement of 3 amino acids especially important to poultry nutrition, cystine, methionine, and lysine, in a single preoxidized hydrolysate; this method can be extended to another 9 protein amino acids.     

非离子表面活性剂PEG对棉秆木质纤维素酶解的影响

为了提高纤维素的水解作用,减少酶的用量,该文研究不同分子量的聚乙二醇(PEG,poly ethylene glycol)表面活性剂对棉秆木质纤维素水解复合酶的影响,研究不同分子量的PEG对还原糖、纤维素酶、木质素酶、蛋白吸附率、酶动力学特征的影响。结果表明,2.5g/L添加量的PEG 2000、PEG 4000、PEG 6000和PEG 8000均能提高棉秆木质纤维素水解酶的活力,增加纤维素的转化率,减少蛋白的非特异性吸附率,提高棉秆木质素和纤维素的降解率,提高最大反应速度和降低酶与底物的亲和力,其中PEG 6000效果最好。3.0g/L的PEG 6000添加量对纤维素的转化率最高,达到58.12%(P<0.05)。对于吸附率,2.0、3.0、4.0、5.0g/L的添加量与对照相比差异显著(P<0.05),且相互之间差异不显著(P>0.05)。添加3.0g/L PEG 6000的表面活性剂能保持上清液中较高的蛋白浓度,水解3h后,没有添加表面活性剂的处理,纤维滤纸酶酶活从0.1245FPU/mL下降到0.012FPU/mL,下降了90.37%,添加PEG 6000后,FPA酶的活性恢复到0.041FPU/mL;水解48h,添加PEG 6000和不添加PEG 6000纤维素的转化率分别达到65.71%和54.68%,添加PEG 6000提高纤维素转化率20.18%。结果表明,PEG6000可以提高纤维素的转化率,为工业生产实践提供了参考。     

Some sulfur-containing metabolites of tri-n-butyltin chloride in male rats.

In an attempt to elucidate metabolic destination of TBTO, sulfur-containing metabolites were investigated in the urine. Tri-n-butyltin chloride (TBTC), tri-n-butyltin oxide (TBTO), and their in vitro metabolites in rat liver microsomal enzyme systems, di-n-butyl(3-hydroxybutyl)tin chloride (T3OH), di-n-butyl(3-oxobutyl)tin chloride (T3CO), dibutyltin dichloride (DBTC), and monobutyltin trichloride (MBTC), were intraperitoneally administered to rats. In particular, administration of T3OH and T3CO gave higher amounts of mercapturic acid derivatives, such as N-acetyl-S-(3-oxobutyl)-L-cysteine (3CO-MA) and N-acetyl-S-(3-hydroxybutyl)-L-cysteine (3OH-MA), than TBTC or TBTO. On the other hand, DBTC and MBTC did not yield measurable amounts of 3CO-MA and/or 3OH-MA. The appearance of organotin metabolites in urine indicates that T3OH, T3CO, and hypothesized secondary metabolites, such as n-butyl(3-hydroxybutyl)(3-oxobutyl)tin chloride, n-butyl(3-hydroxybutyl)(4-hydroxybutyl)tin chloride, etc., are subject to the action of glutathione S-transferase to give mercapturic acid derivatives. These sulfur-containing metabolites (3CO-MA and 3OH-MA) were also found in control rat urine.     

Identification of character impact odorants of different soybean lecithins.

The potent odorants of standardized, enzymatically hydrolyzed, and deoiled soybean lecithins were characterized systematically by combined gas chromatography/mass spectrometry and olfactometry. Sixty-one odorants were identified; 53 of these odor-active compounds have not previously been reported as odorants of soybean lecithin flavor. By aroma extract dilution analysis and modified combined hedonic and response measurement the following odorants showed the highest flavor dilution factors and CHARM values: (E,E)-2, 4-decadienal (deep-fried), (E)-beta-damascenone (apple-like), 2, 3-diethyl-5-methylpyrazine (roasty, earthy), (E)-2-nonenal (cardboard-like), trans-4,5-epoxy-(E)-2-decenal (metallic), 1-nonen-3-one (mushroom-like), 2-ethyl-3,5-dimethylpyrazine (roasty, earthy), and 1-octen-3-one (mushroom-like). Enzymatic hydrolysis intensified especially the roasty sensation of 2, 3-diethyl-5-methylpyrazine, whereas deoiling effected a general significant decrease in olfactory perception on the nitrogen-containing compounds. In addition, sensory profiles of nasal and retronasal lecithin odor were performed.