Enzymatic lipophilization of epicatechin with free fatty acids and its effect on antioxidative capacity in crude camellia seed oil.

BACKGROUND: Crude camellia seed oil is rich in free fatty acids, which must be removed to produce an oil of acceptable quality. In the present study, we reduced the free fatty acid content of crude camellia seed oil by lipophilization of epicatechin with these free fatty acids in the presence of Candida antarctica lipase B (Novozym 435), and this may enhance the oxidative stability of the oil at the same time. RESULTS: The acid value of crude camellia seed oil reduced from 3.7 to 2.5 mgKOH g-1 after lipophilization. Gas chomatography-mass spectrometry analysis revealed that epicatechin oleate and epicatechin palmitate were synthesized in the lipophilized oil. The peroxide, p-anisidine, and total oxidation values during heating of the lipophilized oil were much lower than that of the crude oil and commercially available camellia seed oil, suggesting that lipophilized epicatechin derivatives could help enhance the oxidative stability of edible oil. CONCLUSION: The enzymatic process to lipophilize epicatechin with the free fatty acids in crude camellia seed oil described in the present study could decrease the acid value to meet the quality standards for commercial camellia seed oil and, at the same time, obtain a new edible camellia seed oil product with good oxidative stability. © 2016 Society of Chemical Industry.

Effects of microwave pretreatment and transglutaminase crosslinking on the gelation properties of soybean protein isolate and wheat gluten mixtures.

BACKGROUND: The integration of soybean protein isolate (SPI) with wheat gluten (WG) crosslinked via microbial transglutaminase (MTGase) may enhance the formation of ϵ-(γ-glutamyl)lysine covalent bonds, because SPI is rich in lysine and WG contains more glutamine. Microwave pretreatment may accelerate enzymatic reactions. In this study, we aimed to elucidate the effects of microwave pretreatment on the gelation properties of SPI and WG crosslinked with MTGase. RESULTS: Interestingly, the gel strength, water-holding capacity (WHC) and storage modulus (G') values of MTGase-induced SPI/WG gels were dramatically improved with increasing microwave power. Moreover, the MTGase crosslinking reaction promoted the formation of disulfide bonds, markedly reducing the free SH group and soluble protein content of gels. Fourier transform infrared spectroscopic analysis of SPI/WG gels showed that microwave pretreatment increased the proportion of α-helices and ß-turns and decreased the proportion of ß-sheets. Results from scanning electron microscopy indicated that the MTGase-induced SPI/WG gels had denser and more homogeneous microstructures after microwave pretreatment. CONCLUSION: The effect of microwave pretreatment is useful in advancing gelation characters of MTGase-induced SPI/WG gels and provides the possibility for expanding the application of food protein. © 2015 Society of Chemical Industry.

Cloning, expression, and characterization of a novel xylose reductase from Rhizopus oryzae.

Rhizopus oryzae is valuable as a producer of organic acids via lignocellulose catalysis. R. oryzae metabolizes xylose, which is one component of lignocellulose hydrolysate. In this study, a novel NADPH-dependent xylose reductase gene from R. oryzae AS 3.819 (Roxr) was cloned and expressed in Pichia pastoris GS115. Homology alignment suggested that the 320-residue protein contained domains and active sites belonging to the aldo/keto reductase family. SDS-PAGE demonstrated that the recombinant xylose reductase has a molecular weight of approximately 37 kDa. The optimal catalytic pH and temperature of the purified recombinant protein were 5.8 and 50 °C, respectively. The recombinant protein was stable from pH 4.4 to 6.5 and at temperatures below 42 °C. The recombinant enzyme has bias for D-xylose and L-arabinose as substrates and NADPH as its coenzyme. Real-time quantitative reverse transcription PCR tests suggested that native Roxr expression is regulated by a carbon catabolite repression mechanism. Site-directed mutagenesis at two possible key sites involved in coenzyme binding, Thr(226) → Glu(226) and Val(274) → Asn(274), were performed, respectively. The coenzyme specificity constants of the resulted RoXR(T226E) and RoXR(V274N) for NADH increased 18.2-fold and 2.4-fold, which suggested possibility to improve the NADH preference of this enzyme through genetic modification.

Preparation and evaluation of antioxidant activities of peptides obtained from defatted wheat germ by fermentation.

Defatted wheat germ peptides (DWGPs) were prepared by fermentation with Bacillus Subtilis B1 and the antioxidant activities of DWGPs were investigated. The fermentation condition was optimized by response surface method (RSM) with three factors and three levels according to Box-Behnken theory. A maximal yield of DWGPs was achieved 8.69 mg/mL under optimal conditions: inoculum size 8%, fermentation temperature 31 °C and time 48 h. The main portion in the hydrolysates after fermentation was not free amino acid but peptide. The main molecular weight distribution of DWGPs was lower than 1000 Da. A positive correlation (R(2) = 0.9911) was found between concentration of DWGPs and total antioxidant capacity (T-AOC). DWGPs presented a significant does-dependent on scavenging activities of DPPH, hydroxyl and superoxide anion radicals. The EC50 values for the scavenging rates of DPPH, hydroxyl and superoxide anion radicals were 3.16 mg/mL, 6.04 mg/mL and 7.46 mg/mL, respectively. The results suggested that DWGPs produced by fermentation could be used as a promising antioxidant ingredient.

Fabrication and characterization of soy ß-conglycinin-dextran-polyphenol nanocomplexes: Improvement on the antioxidant activity and sustained-release property of curcumin.

In this study, glycated soy ß-conglycinin (ß-CG) stabilized curcumin (Cur) composites were fabricated by a unique reversible self-assembly character of ß-conglycinin-dextran conjugates (ß-CG-DEX). Intrinsic fluorescence and far-UV CD spectra revealed that glycation did not affect the self-assembly property of ß-CG in the pH-shifting treatment. The structure of ß-CG-DEX could be unfolded at pH 12.0 and reassembled during acidification (from pH 12.0 to 7.0). Meanwhile, ß-CG-DEX-3d, which was incubated at 60 °C for 3 days, exhibited a high loading capacity (123.4 mg/g) for curcumin, which far exceeds that (74.90 mg/g) of ß-CG-Cur. Moreover, the reassembled ß-CG-DEX-3d-Cur showed eminent antioxidant activity of approximately 1.5 times higher than that of free curcumin. During the simulated gastrointestinal condition, compared with ß-CG-Cur, ß-CG-DEX-3d-Cur nanoparticles showed a more stable and sustained release of curcumin. Thus, ß-CG-DEX has immense potential to become a new delivery carrier for hydrophobic food components by means of a self-assembly strategy.

Transcriptional regulation of the gene for prothoracicotropic hormone in the silkworm, Bombyx mori.

Prothoracicotropic hormone (PTTH) is one of key players in regulation of insect growth, molting, metamorphosis, diapause, and is expressed specifically in the two pairs of lateral PTTH-producing neurosecretory cells in the brain. Analysis of cis-regulatory elements of the PTTH promoter might elucidate the regulatory mechanism controlling PTTH expression. In this study, the PTTH gene promoter of Bombyx mori (Bom-PTTH) was cloned and sequenced. The cis-regulatory elements in Bom-PTTH gene promoter were predicted using Matinspector software, including myocyte-specific enhancer factor 2, pre-B-cell leukemia homeobox 1, TATA box, etc. Transient transfection assays using a series of fragments linked to the luciferase reporter gene indicated that the fragment spanning -110 to +33 bp of the Bom-PTTH promoter showed high ability to support reporter gene expression, but the region of +34 to +192 bp and -512 to -111 bp repressed the promoter activity in the BmN and Bm5 cell lines. Electrophoretic mobility shift assays demonstrated that the nuclear protein could specifically bind to the region spanning -124 to -6 bp of the Bom-PTTH promoter. Furthermore, we observed that the nuclear protein could specifically bind to the -59 to -30 bp region of the Bom-PTTH promoter. A classical TATA box, TATATAA, localized at positions -47 to -41 bp, which is a potential site for interaction with TATA box binding protein (TBP). Mutation of this TATA box resulted in no distinct binding band. Taken together, TATA box was involved in regulation of PTTH gene expression in B. mori.

The complete nucleotide sequence of the mitochondrial genome of Phthonandria atrilineata (Lepidoptera: Geometridae).

Using long-polymerase chain reaction (Long-PCR) method, we determined the complete nucleotide sequence of the mitochondrial genome (mitogenome) of Phthonandria atrilineata. The complete mtDNA from P. atrilineata was 15,499 base pairs in length and contained 13 protein-coding genes (PCGs), 2 rRNA genes, 22 tRNA genes, and a control region. The P. atrilineata genes were in the same order and orientation as the completely sequenced mitogenomes of other lepidopteran species. The nucleotide composition of P. atrilineata mitogenome was biased toward A + T nucleotides (81.02%), and the 13 PCGs show different A + T contents that range from 73.25% (cox1) to 92.12% (atp8). Phthonandria had the canonical set of 22 tRNA genes, that fold in the typical cloverleaf structure described for metazoan mt tRNAs, with the unique exception of trnS(AGN). The phylogenetic relationships were reconstructed with the concatenated sequences of the 13 PCGs of the mitochondrial genome, which confirmed that P. atrilineata is most closely related to the superfamily Bombycoidea.

Preparation of camellia oil-based W/O emulsions stabilized by tea polyphenol palmitate: Structuring camellia oil as a potential solid fat replacer.

Water-in-oil (W/O) emulsions with a discontinuous aqueous phase dispersed in a continuous camellia oil phase were prepared by using tea polyphenol palmitate (Tp-palmitate) particle as effective stabilizers and their properties were characterized by droplet size, slip melting point (SMP), stability, microstructure and rheology. The d(4,3) and d(3,2) decreased from 7.96 µm to 4.67 µm and from 5.98 µm to 3.07 µm, respectively, and the SMP rose from 33.73 °C to 38.60 °C when the Tp-palmitate concentration increased from 1.0% to 2.5% (m/v). The storage stability, freeze/thaw stability and thermal stability significantly enhanced and the droplets aggregation progressively increased with the increasing of Tp-palmitate concentration. The liquid camellia oil was transformed into solid-like viscoelastic emulsion gels with a SMP of 38.6 °C when using 2.5% Tp-palmitate as particle stabilizers. This study provides a promising method for production of edible gel-like W/O emulsions using polyphenol-lipid complexes to potentially replace solid fats.

Preparation and properties of OSA-modified taro starches and their application for stabilizing Pickering emulsions.

In this study, the octenylsuccinylated taro starches (OSTS) with different degree of substitution (DS, from 0.009 to 0.032) were prepared and their structural properties such as granule size, wettability and morphology were studied. The purpose of this work was to elucidate the OSTS with different DS using as particle stabilizers for Pickering emulsions, and the effect of DS on the stability, droplet size, microstructure and rheological properties of OSTS-stabilized emulsions were investigated. Octenylsuccinic anhydride (OSA) modification had slight effects on the morphology or granule size of taro starch, but markedly increased the contact angle from 25.4° to 70.1°. Octenylsuccinylation significantly improved the emulsifying capacity of taro starch granules, and thus OSTS-stabilized emulsions formed at higher DS exhibited better stability. Droplet size distribution results and microscopic observations revealed that OSTS-emulsion prepared at DS of 0.032 had the smallest droplet size and most uniform distribution compared with the other emulsions. The rheological results indicated that both OSTS-emulsions (DS, from 0.009 to 0.032) showed shear-thinning behavior as a non-Newtonian fluid, and the viscosities of emulsions were progressively improved with the increase of DS. Moreover, the G' and G″ values of OSTS-emulsions increased with increasing DS, reflecting the enhanced viscoelastic properties and exhibiting an improved rigidity of the emulsions. The above results suggested that higher-DS favored the formation of superior OSTS-emulsions, and thus OSTS with a high DS (DS ≥ 0.018) can be used for preparing stable Pickering emulsions.

Molecular characters and expression analysis of the gene encoding eclosion hormone from the Asian corn borer, Ostrinia furnacalis.

Using rapid amplification of cDNA ends (RACE), the cDNA encoding eclosion hormone (EH) was cloned from the brain of Ostrinia furnacalis. The full Osf-EH cDNA is 986 bp and contains a 267 bp open reading frame encoding an 88 amino acid preprohormone, which including a hydrophobic 26 amino acid signal peptide and a 62 amino acid mature peptide. The mature Osf-EH shows high identity with Manduca sexta (95.2%), Helicoverpa armigera (91.9%) and Bombyx mori (85.5%), but low identify with Tribolium castaneum (63.6%), Drosophila melanogaster (56.5%) and Apis mellifera (54.8%). Using the HMMSTR Prediction Server, the 3D structure of Osf-EH was modeled. There are four beta-turns and three alpha-helixes predicted in Osf-EH, with the pattern of beta-beta-alpha-alpha-beta-beta-alpha. Northern blot analysis indicated a 1.0 kb transcript present only in the brain. The Osf-EH mRNA can not be detected in other neural tissues, such as the suboesophageal ganglion, thoracic ganglion, abdominal ganglion and other non-neural tissues, such as the midgut, fat body and epidermis. The Osf-EH mRNA content in the brain was measured using the combined method of quantitative RT-PCR and Southern blotting, which reached its highest level the day before the molt.

Enhancement of growth and polysaccharide production in suspension cultures of protocorm-like bodies from Dendrobium huoshanense by the addition of putrescine.

Putrescine at 0.6 mM stimulated protocorm-like body growth and polysaccharide synthesis in suspension cultures of Dendrobium huoshanense. The specific growth rate of protocorm-like body increased from 0.047 to 0.056 day(-1), and the maximum dry weight and polysaccharide production reached 33.2 and 2.94 g l(-1), respectively, while they were 24.6 and 2.12 g l(-1), respectively, in the control. The administration of polyamine inhibitor, alpha-DL-difluoromethylarginine, at 1 mM, decreased protocorm-like body growth and polysaccharide production to 21.4 and 1.76 g l(-1), respectively.

Changes in chemical interactions and protein conformation during heat-induced wheat gluten gel formation.

In order to elucidate the heat-induced wheat gluten gel formation mechanism, changes in chemical interactions and protein conformation were investigated during gelation. The contribution of ionic and hydrogen bonds were found to decrease from 0.746 and 4.133g/L to 0.397 and 2.733g/L, respectively, as the temperature increased from 25 to 90°C. Moreover, the free SH content remarkably decreased from 37.91 to 19.79µmol/g during gelation. Ultraviolet absorption spectra and intrinsic fluorescence spectra suggested that wheat gluten unfolded during the heating process. In addition, wheat gluten gels treated at 80 and 90°C exhibited a "steric hindrance" effect, which can be attributed to the formation of aggregates. Fourier transform infrared spectra suggested that the random coil content increased at low temperatures (40 and 50°C), whereas the content of intermolecular ß-sheets due to protein aggregation increased from 38.10% to 44.28% when the gelation temperature was 90°C.

Transglutaminase-set colloidal properties of wheat gluten with ultrasound pretreatments.

The low solubility of wheat gluten limits its accessibility. This work aimed to study the impact of ultrasonic pretreatments on the gelation of wheat gluten. The pretreatments included ultrasound combined with alkali, urea, Na2SO3, with or without the addition of transglutaminase (TGase). The gel strength of wheat gluten was 287g/cm2 after treatment with Na2SO3/ultrasound/TGase. The free sulfhydryl and disulfide bond content was significantly affected by ultrasound treatment. After treatments including TGase crosslinking, the molecular weight of wheat gluten complexes became larger. The network formed by the wheat gluten was transformed into a dense and homogenous structure after the pretreatment with Na2SO3/ultrasound/TGase. The content of random coil of wheat gluten increased. The gelation of wheat gluten could also be significantly enhanced by Na2SO3/ultrasound treatment followed by TGase treatment. Using physical and chemical pretreatments to allow TGase to enhance the gelation of wheat gluten may increase its uses as a food additive.

Species diversity of Lachnum (Helotiales, Hyaloscyphaceae) from temperate China.

Twenty-three temperate China species of Lachnum, Lachnum abnorme, L. angustum, L. brevipilosum, L. calosporum, L. calyculiforme, L. carneolum, L. ciliare, L. controversum, L. flavidulum, L. cf. fushanese, L. indicum, L. kumaonicum, L. lushanese, L. minutum, L. montanum, L. cf. pteridophyllum, L. pygmaeum, L. sclerotii var. sclerotii, L. sclerotii var. sichuanense, L. subpygmeaum, L. tenuissimum, L. virgineum and L. willisii are reported, whose main characteristics are given in a formula of the described species, some of which are discussed below.

[Screening of a low alcohol dehydrogenase activity mutant of rhizopus oryzae and the regulation of Zn2+ and Mg2+].

Ethanol is the main by-product in the fermentation broth of Rhizopus oryzae As3.3461 for the production of high-optical purity L-lactic acid. Alcohol Dehydrogenase (ADH) is the branch pathway enzyme that catalyzes the transformation of ethanol from pyruvate in Rhizopus oryzae, which decreases the conversion rate of glucose to L-lactic acid. Thus, screening the mutants with lower ADH activity may increase lactate production dramatically. In present study, Rhizopus oryzae As3.3461 was mutated with N-methyl-N'-nitro-N-nitrosoguanidine (NTG), and 21 mutants which showed lower ADH activity were isolated with selective medium of Yeast-Peptone-Dextrose (YPD) containing 0.6% allyl alcohol (V/V). Compared with other mutants, the 12th mutant strain (named as HBF-12) shows the highest conversion rate of L-lactic acid. By contrast with Rhizopus oryzae As3.3461, the parent strain, the ethanol production and the ADH activity of HBF-12 decrease 73.6% and 76%, respectively. Whereas, the L-lactic acid production and the LDH activity of HBF-12 increase 41.2% and 19.6% than those of the parent strain, respectively. The activities of ADH and LDH of HBF-12 were regulated by Zn2+ and Mg2+, but showed opposite effects. Added with Zn2+ to the concentration of 0.01% improves the ADH activity dramatically, but inhibits the activity of LDH. By contraries, added with Mg2+ improves the LDH activity markedly, but inhibits the ADH activity slightly. In fermentation experiment, the addition of Zn2+ and Mg2+ show different effects on the accumulation of ethanol, L-lactic acid and the biomass in mutant HBF-12. When improve the concentration of Zn2+, the accumulation of L-lactic acid and the biomass show the decreased trend, but the production of ethanol show positive effect. With the improvement of the concentration of Mg2+, the production of lactic acid and biomass increase markedly, but no effect on the production of ethanol. When ferment under the concentrations of Zn2+ 0.01% and Mg2+ 0.04% in fermentation medium, the lactate production of HBF-12 reached the highest level, 96.21 g/L.

Transglutaminase-induced gelation properties of soy protein isolate and wheat gluten mixtures with high intensity ultrasonic pretreatment.

Soy protein isolate (SPI) and wheat gluten (WG) are widely used in commercial food applications in Asia for their nutritional value and functional properties. However, individually each exhibits poor gelation. In this study, we examined the microbial transglutaminase (MTGase)-induced gelation properties of SPI and WG mixtures with high intensity ultrasonic pretreatment. Ultrasonic treatment reduced the particle size of SPI/WG molecules, which led to improvements in surface hydrophobicity (Ho) and free sulfhydryl (SH) group content. However, MTGase crosslinking facilitated the formation of disulfide bonds, markedly decreasing the content of free SH groups. Ultrasonic treatment improved the gel strength, water holding capacity, and storage modulus and resulted in denser and more homogeneous networks of MTGase-induced SPI/WG gels. In addition, ultrasonic treatment changed the secondary structure of the gel samples as determined by Fourier transform infrared spectroscopic analysis, with a reduction in α-helices and ß-turns and an increase in ß-sheets and random coils. Thus, ultrasound is useful in facilitating the gelation properties of MTGase-induced SPI/WG gels and might expand their utilization in the food protein gelation industry.

Effect of Modified Wheat Gluten on Boiling Resistance Capacity of Pork Meatballs.

The effect of the modified wheat gluten (MWG) extender, prepared by alcalase-based hydrolysis and transglutaminase cross-linking, on meatballs was analyzed in this study. Here, we studied the effect of MWG addition on the boiling resistance capacity of pork meatballs (MB-MWG) at high temperature (100 °C) and increasing cooking time; meatballs with added soy protein isolates (MB-SPI) and raw wheat gluten (MB-WG) were used as references. The cooking loss, water-holding capacity (WHC), and textural properties of meatballs were investigated. The results revealed that MB-MWG showed lower cooking loss, which decreased by 49.16% compared to meatballs without added extenders when treated for 30 min. The WHC of MB-MWG significantly increased from 80.68% to 95.42%. The hardness, springiness, and chewiness (textural properties) of MB-MWG were also significantly increased by 97.05%, 6.68%, and 121.96%, respectively. The addition of MWG increased the cross-linking in meatballs during the cooking process, as indicated by the higher G'. SDS-PAGE indicated an obvious decrease in myosin heavy chain in MB-MWG cooked for 30 min at 100 °C, which was attributed to the interaction of myofibrillar proteins in pork meat with MWG. The nuclear magnetic resonance T2 relaxation time patterns indicated that MWG addition caused an increase in the bound water content, and decrease in the free water content, of meatballs. An analysis of the microstructures revealed that the MB-MWG formed the most regular and compact network. Therefore, MWG could be used as an ingredient to facilitate the processing of meat products.

[Partial least-squares regression analysis for simultaneous spectrophotometric determination of Fe3+ and Al3+].

Adding hexodecyltrimethylammonium bromide(CTMAB) to improve solubility and delicacy, and using chromazurol S as a chromogenic reagent, and acetone as a stabilizing reagent, partial least-squares regression(PLS) was used to determine simultaneously trace amounts of Fe(III) and Al(III) in sythetic samples. Considering the stability of the rays, and the absorption properties of the two ingredients, seven wavelengths between 610 and 670 nm were selected for data collection. As a comparison, the methods of CPA and PLS were used respectively for analysing the data obtained from the experiment, and the results suggested that the latter was much better.

[Callus formation and its isoflavonoid accumulation in Maackia amurensis].

OBJECTIVE: To obtain Maackia amurensis callus and investigate some factors influencing total isoflavonoid production in callus. METHOD: Different media were used for callus induction from cotyledonary and hypocotyl explants. Total isoflavonoid content was determined by UV spectrophotometer. RESULT: Medium types and hormone combinations influenced both callus formation and their developmental states. MS medium supplemented with NAA/BA or containing 2,4-D was suitable for callus induction from cotyledonary explants. Among the basal media tested, MS, N6 and B5 were suitable for cotyledonary callus formation and SH for hypocotyl callus formation. The formed callus could synthesize isoflavones. Media containing NAA were suitable for isoflavonoid production in cotyledonary callus and media containing 2,4-D/BA were suitable for hypocotyl callus, but 2,4-D/KT at high concentration inhibited the isoflavonoid accumulation in hypocotyl and coytledonary calli. According to the developmentally morphological characters, the formed callus could be classified into four types. II-type callus with yellow in color and friability in texture showed the highest accumulation of isoflavones. CONCLUSION: The influences of medium type and hormone combinations on Maackia amurensis callus formation are reflected in both developmentally morphological characters and isoflavonoid accumulation of calli. Yellow, friable callus induced from cotyledonary and hypocotyl explants in MS or N6 medium supplemented with 2,4-D and BA exhibit the optimum growth and isoflavonoid production.

[Suspension culture of protocorm-like bodies from the endangered medicinal plant Dendrobium huoshanenese].

OBJECTIVE: To investigate the characteristics of growth, and water-soluble polysaccharide and total alkaloid accumulation in protocom-like bodies (PLBs) of Dendrobium huoshanenese in liquid culture system. METHOD: PLBs were suspended in liquid medium and growth kinetics was analyzed. Water-soluble polysaccharide and total alkaloid content in PLBs were determined by colorimetry. RESULT: PLBs were induced from stem explants of D. huoshanenese regenerants cultured on MS medium supplemented with NAA or NAA and KT at different concentrations. Basal MS medium was suitable for propagation of PLBs. When PLBs were suspended in liquid medium, mumax was 0.044.d-1, t(d) was 15.8 d and the optimum growth time was 4 weeks. Water-soluble polysaccharide and total alkaloid contents in PLBs were 3.75% and 0.0261%, respectively. CONCLUSION: PLBs in liquid culture system show a potential for rapid growth and high metabolite synthesis, which provides possibility for exploiting resources of D. huoshanenese by large scale culture of PLBs.