Inhibitory effect of carvacrol against Alternaria alternata causing goji fruit rot by disrupting the integrity and composition of cell wall.

Goji (Lycium barbarum L.) is a widely planted crop in China that is easily infected by the pathogenic fungus Alternaria alternata, which causes rot after harvest. Previous studies showed that carvacrol (CVR) significantly inhibited the mycelial growth of A. alternata in vitro and reduced Alternaria rot in goji fruits in vivo. The present study aimed to explore the antifungal mechanism of CVR against A. alternata. Optical microscopy and calcofluor white (CFW) fluorescence observations showed that CVR affected the cell wall of A. alternata. CVR treatment affected the integrity of the cell wall and the content of substances in the cell wall as measured by alkaline phosphatase (AKP) activity, Fourier transform-infrared spectroscopy (FT-IR), and X-ray photoelectron spectroscopy (XPS). Chitin and ß-1,3-glucan contents in cells decreased after CVR treatment, and the activities of ß-glucan synthase and chitin synthase decreased. Transcriptome analysis revealed that CVR treatment affected cell wall-related genes in A. alternata, thereby affecting cell wall growth. Cell wall resistance also decreased with CVR treatment. Collectively, these results suggest that CVR may exert antifungal activity by interfering with cell wall construction, leading to impairment of cell wall permeability and integrity.

Molecular characteristics and physicochemical properties of very small granule starch isolated from Agriophyllum squarrosum seeds.

Novel starch resources isolated from accessible botanical origins are of special interest to food scientists in the context of food security. In this study, Agriophyllum squarrosum starches (AS-1, AS-2, and AS-3) were isolated from three ecotypes of A. squarrosum seeds and compared with quinoa starch (QS). The mean particle diameter of AS granules ranged from 1.12 to 1.15 µm, and AS amylopectin had a significantly higher Mw than QS (p < 0.05). Compared with QS, AS samples had more branching and substitution of amylopectin structures. The peak viscosity, breakdown viscosity, and swelling degree of the AS samples were significantly lower than those of QS (p < 0.05). AS showed a lower crystalline degree and higher gelatinization temperatures, and the freshly cooked AS showed a slower digestibility rate than QS. The physicochemical properties and chain profiles of AS facilitate the application of AS and the domestication of A. squarrosum crops.

Effects of exogenous α-amylases, glucoamylases, and proteases on ruminal in vitro dry matter and starch digestibility, gas production, and volatile fatty acids of mature dent corn grain.

Two separate experiments were carried out to evaluate the effects of incremental doses of 10 exogenous endo-acting α-amylase and exo-acting glucoamylase; 1LAT (bacterial α-amylase), 2AK, 3AC, 4Cs4, 5Trga, 6Afuga, 7Fvga, and 10Tg (fungal α-amylases, glucoamylases, and α-glucosidase), 8Star and 9Syn (fungal amylase-mixtures; experiment 1) and three exogenous proteases; 11P14L, 12P7L, and 13P30L (bacterial proteases; experiment 2) on in vitro dry matter digestibility (IVDMD) and in vitro starch digestibility (IVSD) of mature dent corn grain using a batch culture system. Incremental doses of the exogenous enzymes (0, 0.25, 0.50, 0.75, and 1.00 mg/g of dried substrate) were applied directly to the substrate (0.5 g of ground corn, 4 mm) in sextuplicate (experiment 1) or quadruplicate (experiment 2) within F57 filter bags, which were incubated at 39 °C in buffered rumen fluid for 7 h. Rumen fluid was collected 2-3 h after the morning feeding from three lactating dairy cows and pooled. Cows were consuming a midlactation total mixed ration (TMR; 1.60 Mcal/kg DM and 15.4%; net energy of lactation and crude protein, respectively). Three independent runs were carried out for each experiment. Data were analyzed as a randomized complete block design using run as the blocking factor. Dose was used as a fixed factor while run was considered a random factor. Linear, quadratic, and cubic orthogonal contrasts were also tested. In experiment 1, enzymes 2AK, 3AC, and 10Tg did not increase (P > 0.10) IVDMD and IVSD, whereas 0.25 mg of enzymes 1LAT, 5Trga, and 8Star increased (P < 0.01) IVDMD by 23%, 47%, and 62% and IVSD by 35%, 41%, and 58%, respectively, compared with the control. Enzymes 4Cs4, 6Afuga, 7Fvga, and 9Syn linearly increased IVDMD and IVSD (P < 0.01). Greatest increases in IVDMD (82.9%) and IVSD (85.9%) resulted with 1 mg of 6Afuga compared to control. In experiment 2, the lowest dose of exogenous proteases 11P14L and 12P7L increased (P < 0.01) IVDMD by 98% and 87% and IVSD by 57% and 64%, respectively, whereas the highest dose of 13P30L increased (P = 0.02) IVDMD by 44.8% and IVSD by 30%, relative to the control. In conclusion, IVSD and IVDMD were increased by one α-amylase, certain glucoamylases, and all proteases tested, with the glucoamylase 6Afuga in experiment 1 and the neutral protease 12P7L in experiment 2, increasing IVDMD and IVSD to the greater extents. Future in vivo studies are required to validate these findings before these enzyme additives can be recommended for improving the digestibility of mature dent corn grain.

Effect of exogenous glucoamylase inclusion on in vitro fermentation and growth performance of feedlot steers fed a dry-rolled corn-based diet.

The objective was to evaluate the effect of adding an exogenous glucoamylase (GA) enzyme from the fungus Trichoderma reesei on in vitro fermentation, growth performance, and carcass characteristics of feedlot steers fed a dry-rolled corn (DRC)-based diet. Experiment 1 evaluated three levels of added enzyme (0, 0.24, and 0.72 GA enzyme units) and two corn particle sizes (CPS; 2 and 4 mm) in a factorial arrangement using a 7 h in vitro batch culture fermentation. Addition of GA increased (P < 0.01) in vitro dry matter disappearance by 13% and decreased final pH (P < 0.01). Molar proportion of propionate increased with GA inclusion (P < 0.01). A smaller CPS increased (P < 0.01) in vitro dry matter disappearance and total volatile fatty acid and decreased final pH (P < 0.01). A smaller CPS also decreased (P < 0.01) the molar proportion of acetate and increased (P < 0.01) the molar proportion of butyrate. In experiment 2, Angus × Simmental steers (n = 105; initial body weight [BW] = 329 ± 38 kg) were used to evaluate the inclusion of an exogenous GA on growth performance and carcass characteristics. Steers were fed a basal diet consisting of 60% DRC, 17.5% modified distillers grains with solubles, 12.5% corn silage, and 10% dry supplement on a dry matter basis for 136 d. Steers were blocked by weight and allotted to pens. Pens were randomly assigned to one of three treatments (5 pens/treatment): diet with no GA (CON), low inclusion of GA (122 enzyme units/kg dry matter [DM]; LGA), or high inclusion of GA (183 enzyme units/kg DM; HGA). Inclusion of GA did not affect (P ≥ 0.23) final BW, dry matter intake (DMI), or average daily gain (ADG) for the 136-d feeding period. Feed conversion was affected (P = 0.02) by treatment with steers fed HGA having ~8% greater G:F compared with LGA and CON. Treatment did not affect (P = 0.32) fecal starch. Inclusion of GA did not affect (P ≥ 0.19) carcass traits including hot carcass weight, 12th rib fat thickness, yield grade, longissimus muscle area, or marbling score. Overall, results suggest inclusion of exogenous GA enzyme increased in vitro dry matter disappearance in batch culture and improved feed conversion in steers fed 183 enzyme units/kg DM during the finishing phase.

Inhibition of preadipocyte differentiation by Lycium barbarum polysaccharide treatment in 3T3-L1 cultures

BACKGROUND: Lycium barbarum (also called wolfberry), a famous Chinese traditional medicine and food ingredient, is well recognized for its significant role in preventing obesity; however, the molecular mechanisms underlying its preventive effects on fat accumulation are not well understood yet. The aim of this study was to determine the effects and mechanism of Lycium barbarum polysaccharides (LBP) on the proliferation and differentiation of 3T3-L1 preadipocytes. MTT was used to detect the proliferation of 3T3-Ll preadipocytes. Oil red O staining and colorimetric analysis were used to detect cytosolic lipid accumulation during 3T3-L1 preadipocyte differentiation. Real-time fluorescent quantitative PCR (qPCR) technology was used to detect peroxisome proliferator-activated receptor c (PPARc), CCAAT/enhancer-binding protein a (C/EBPa), adipocyte fatty-acid-binding protein (aP2), fatty acid synthase (FAS), and lipoprotein lipase (LPL) expression. RESULTS: The concentration of LBP from 25 to 200 lg/mL showed a tendency to inhibit the growth of preadipocytes at 24 h, and it inhibited the differentiation of 3T3-L1 preadipocytes in a dose-dependent manner. In the preadipocytes treated with 200 lg/mL LBP, there were reduced lipid droplets in the cytoplasm, and its effect was opposite to that of rosiglitazone (ROS), which significantly reduced the PPARc, C/EBPa, aP2, FAS, and LPL mRNA expression of adipocytes. CONCLUSIONS: LBP exerts inhibitive effects on the proliferation and differentiation of 3T3-L1 preadipocytes and decreases the cytoplasm accumulation of lipid droplets during induced differentiation of preadipocytes toward mature cells. Above phenomenon might link to lowered expression of PPARc, C/EBPa, aP2, FAS, and LPL after LBP treatment. Thus, LBP could serve as a potential plant extract to treat human obesity or improve farm animal carcass quality via adjusting lipid metabolism.

Comparative study of protease hydrolysis reaction demonstrating Normalized Peptide Bond Cleavage Frequency and Protease Substrate Broadness Index.

Two commercial proteases (subtilisin-typed FNA from Bacillus amyloliquefaciens, and chymotrypsin-like NPP from Nocardiopsis prasina), porcine pepsin, porcine pancreatin having protease activity and their combinations were studied in vitro by LC-MS for their ability to digest soy protein isolate (SPI) under conditions close to those found in the stomach (pH 3.7) and small intestine (pH 6.5). The total number of peptides generated, and their size distribution were obtained under each set of the digestion conditions. These peptides were grouped according to their C-terminal amino acid (AA) residue (P1) and mass, based on which two concepts were proposed, i.e., Normalized Peptide Bond Cleavage Frequency (NPBCF) and Protease Substrate Broadness Index (PSBI). At pH 3.7, FNA+pepsin increased PSBI vs. pepsin alone by 2.7 and 4.9 percentage points (p.p.) at a SPI:protease ratio of 20:1 and 100:1, respectively. At pH 6.5, FNA+pancreatin improved PSBI by 9.1 and 10.2 p.p. at SPI:protease 20:1 and 100:1, respectively, vs. pancreatin alone. NPP generated 38% more peptides than FNA when administered with pancreatin at SPI:protease 200:1:1 and pH 6.5, but FNA alone (28.9) or FNA+pancreatin (29.1) gave a higher PSBI than pancreatin (22.2), NPP (20.3) and NPP+pancreatin (22.0). At pH 3.7 FNA generated 59% and 39% of peptides of pepsin at SPI:protease of 20:1 and 100:1, respectively, and both groups of peptides had similar size distribution. At pH 6.5 more small sized peptides were generated by FNA or FNA+pancreatin than pancreatin and NPP alone or pancreatin+NPP. In conclusion, FNA showed complementary effects with pepsin and pancreatin in terms of PSBI and generated more small sized peptides compared to NPP.

Effects of Exogenous Glucoamylase Enzymes Alone or in Combination with a Neutral Protease on Apparent Total Tract Digestibility and Feces D-Lactate in Crossbred Angus Bulls Fed a Ration Rich in Rolled Corn.

The aim of this study was to evaluate the effect of two glucoamylases (GA) and the combination of one GA with a neutral protease on apparent total tract digestibility in beef bulls fed a total mixed ration (TMR) rich in rolled corn. Sixteen Angus beef bulls (266 ± 4.9 kg of initial BW, and 182 ± 1.7 d of age) were distributed in 4 blocks, each block consisted of 4 animals balanced by BW. The experimental design was a 4 × 4 Latin square (4 blocks and 4 periods, 2 w per period). Four treatments were tested; (1) control, (2) GA preparation from Trichoderma reesei (TrGA); (3) GA from Aspergillus fumigatus (AfuGA); (4) AfuGA in combination with a neutral protease from Bacillus amyloliquefaciens (BamPro). Apparent total tract digestibility and fecal D-lactate concentration were analyzed. Enzyme supplementation, regardless of enzyme type, increased apparent total tract digestibility of dry matter (from 66.7% to 73.1% ± 2.01), and starch (from 74.7% to 81.8% ± 2.25), without affecting feces D-lactate concentration. Irrespective of glucoamylase type, glucoamylase supplementation improved apparent digestibility of dry matter and starch, and the addition of a protease did not have additional benefits on nutrient digestibility.

Fabrication of MnOx-CeO2/cordierite catalysts doped with FeOx and CuO for preferable catalytic oxidation of chlorobenzene.

A series of MnOx-CeO2 catalysts with MOx doping (M = Cu, Fe, Co and La) supported on cordierite were synthesized by the citric acid complex method, showing preferable catalytic oxidation of chlorobenzene. The distribution of active oxides, surface areas, as well as the structural morphology of M-MnOx-CeO2 catalysts varied with the different Mn/Ce and M/Mn molar ratios. Meanwhile, physicochemical properties of these catalysts were characterized by XRD, BET, SEM, TEM, H2-TPR and IR. More importantly, the catalytic oxidation routes were also investigated where the process was from chlorobenzene to CO2, H2O, HCl and other by-products for the FeOx-MnOx-CeO2 and CuO-MnOx-CeO2 catalysts. The CuO-MnOx-CeO2 catalysts showed a higher chlorobenzene conversion, and the measured light-off temperature T90 was approximately 400°C. However, a large amount of chloropropane as main by-products was observed. For the FeOx-MnOx-CeO2 catalysts, more carbon monoxide could be found with inadequate oxidation. Comparative analyses of two catalysts indicated that the main cause of the oxidation activities and mechanisms were different in the oxidation capacity and water absorbability of FeOx and CuO. Nevertheless, all of these catalysts did not exhibit any deactivation due to chloride with a high reaction temperature, with chloride transformed to form HCl in the off-gas stream.

Crystal structure of quinone-dependent alcohol dehydrogenase from Pseudogluconobacter saccharoketogenes. A versatile dehydrogenase oxidizing alcohols and carbohydrates.

The quinone-dependent alcohol dehydrogenase (PQQ-ADH, E.C. 1.1.5.2) from the Gram-negative bacterium Pseudogluconobacter saccharoketogenes IFO 14464 oxidizes primary alcohols (e.g. ethanol, butanol), secondary alcohols (monosaccharides), as well as aldehydes, polysaccharides, and cyclodextrins. The recombinant protein, expressed in Pichia pastoris, was crystallized, and three-dimensional (3D) structures of the native form, with PQQ and a Ca(2+) ion, and of the enzyme in complex with a Zn(2+) ion and a bound substrate mimic were determined at 1.72 Å and 1.84 Å resolution, respectively. PQQ-ADH displays an eight-bladed ß-propeller fold, characteristic of Type I quinone-dependent methanol dehydrogenases. However, three of the four ligands of the Ca(2+) ion differ from those of related dehydrogenases and they come from different parts of the polypeptide chain. These differences result in a more open, easily accessible active site, which explains why PQQ-ADH can oxidize a broad range of substrates. The bound substrate mimic suggests Asp333 as the catalytic base. Remarkably, no vicinal disulfide bridge is present near the PQQ, which in other PQQ-dependent alcohol dehydrogenases has been proposed to be necessary for electron transfer. Instead an associated cytochrome c can approach the PQQ for direct electron transfer.

Super High Dosing with a Novel Buttiauxella Phytase Continuously Improves Growth Performance, Nutrient Digestibility, and Mineral Status of Weaned Pigs.

This study was conducted to evaluate the efficacy of a novel Buttiauxella phytase to pigs fed P-deficient, corn-soybean meal diets. One hundred and twenty crossbred piglets (9.53 ± 0.84 kg) were allocated to one of five treatments which consisted of four low P diets (0.61 % Ca and 0.46 % total P) supplemented with 0, 500, 1,000, or 20,000 FTU/kg phytase as well as a positive control diet (0.77 % Ca and 0.62 % total P). Each treatment had six replicated pens with four pigs per pen. Pigs were fed the experimental diets for 28 days. Phytase supplementation linearly improved (P < 0.05) average daily gain (ADG), feed conversion ratio (FCR), and apparent total tract digestibility (ATTD) of dry matter, gross energy, crude protein, Ca, and P in weaned pigs. Super high dosing with phytase (20,000 FTU/kg) further increased (P < 0.05) ADG compared with 500 FTU/kg phytase inclusion group, as well as ATTD of Ca and P. Metacarpal bone characteristics and several trace mineral concentration in bone, plasma, or organ tissues were linearly (P < 0.05) improved at increasing dose of phytase. Super high dosing with phytase (20,000 FTU/kg) supplementation improved (P < 0.05) Mn and Zn concentration in bone compared to normal dose of phytase supplementation (500 or 1,000 FTU/kg). In conclusion, supplementation of 500 FTU of Buttiauxella phytase/kg and above effectively hydrolyzed phytate in a low-P corn-soybean diet for pigs. In addition, a super high dosing with phytase (20,000 FTU/kg) improved macro- or micro mineral availability and growth performance.

Ferulic acid dehydrodimer and dehydrotrimer profiles of distiller's dried grains with solubles from different cereal species.

Ferulic acid dehydrodimers (DFA) and dehydrotrimers (TriFA) ester-linked to plant cell wall polymers may cross-link not only cell wall polysaccharides but also other cell wall components including proteins and lignin, thus enhancing the rigidity and potentially affecting the enzymatic degradation of the plant cell wall. Corn, wheat, and mixed-cereal distiller's dried grains with solubles (DDGS) were investigated for composition of DFAs and TriFAs by reversed phase high-performance liquid chromatography with ultraviolet detection. Corn DDGS contained 5.3 and 5.9 times higher contents of total DFAs than wheat and mixed-cereal DDGS, respectively. Furthermore, the contents of total TriFAs were 5.7 and 6.3 times higher in corn DDGS than in wheat and mixed-cereal DDGS, respectively. In addition, both corn grains and corresponding DDGS had similar profiles of individual DFAs and TriFAs, indicating that ferulic acid cross-links in the corn cell wall are presumably not modified during fermentation and DDGS processing.

Structural and functional characterization of ochratoxinase, a novel mycotoxin-degrading enzyme.

Ochratoxin, with ochratoxin A as the dominant form, is one of the five major mycotoxins most harmful to humans and animals. It is produced by Aspergillus and Penicillium species and occurs in a wide range of agricultural products. Detoxification of contaminated food is a challenging health issue. In the present paper we report the identification, characterization and crystal structure (at 2.2 Å) of a novel microbial ochratoxinase from Aspergillus niger. A putative amidase gene encoding a 480 amino acid polypeptide was cloned and homologously expressed in A. niger. The recombinant protein is N-terminally truncated, thermostable, has optimal activity at pH ~6 and 66°C, and is more efficient in ochratoxin A hydrolysis than carboxypeptidase A and Y, the two previously known enzymes capable of degrading this mycotoxin. The subunit of the homo-octameric enzyme folds into a two-domain structure characteristic of a metal dependent amidohydrolase, with a twisted TIM (triosephosphateisomerase)-barrel and a smaller ß-sandwich domain. The active site contains an aspartate residue for acid-base catalysis, and a carboxylated lysine and four histidine residues for binding of a binuclear metal centre.

Crystal structure of α-1,4-glucan lyase, a unique glycoside hydrolase family member with a novel catalytic mechanism.

α-1,4-Glucan lyase (EC 4.2.2.13) from the red seaweed Gracilariopsis lemaneiformis cleaves α-1,4-glucosidic linkages in glycogen, starch, and malto-oligosaccharides, yielding the keto-monosaccharide 1,5-anhydro-D-fructose. The enzyme belongs to glycoside hydrolase family 31 (GH31) but degrades starch via an elimination reaction instead of hydrolysis. The crystal structure shows that the enzyme, like GH31 hydrolases, contains a (ß/α)8-barrel catalytic domain with B and B' subdomains, an N-terminal domain N, and the C-terminal domains C and D. The N-terminal domain N of the lyase was found to bind a trisaccharide. Complexes of the enzyme with acarbose and 1-dexoynojirimycin and two different covalent glycosyl-enzyme intermediates obtained with fluorinated sugar analogues show that, like GH31 hydrolases, the aspartic acid residues Asp(553) and Asp(665) are the catalytic nucleophile and acid, respectively. However, as a unique feature, the catalytic nucleophile is in a position to act also as a base that abstracts a proton from the C2 carbon atom of the covalently bound subsite -1 glucosyl residue, thus explaining the unique lyase activity of the enzyme. One Glu to Val mutation in the active site of the homologous α-glucosidase from Sulfolobus solfataricus resulted in a shift from hydrolytic to lyase activity, demonstrating that a subtle amino acid difference can promote lyase activity in a GH31 hydrolase.

A novel metabolic pathway for glucose production mediated by α-glucosidase-catalyzed conversion of 1,5-anhydrofructose.

α-Glucosidase is in the glycoside hydrolase family 13 (13AG) and 31 (31AG). Only 31AGs can hydrate the D-glucal double bond to form α-2-deoxyglucose. Because 1,5-anhydrofructose (AF), having a 2-OH group, mimics the oxocarbenium ion transition state, AF may be a substrate for α-glucosidases. α-Glucosidase-catalyzed hydration produced α-glucose from AF, which plateaued with time. Combined reaction with α-1,4-glucan lyase and 13AG eliminated the plateau. Aspergillus niger α-glucosidase (31AG), which is stable in organic solvent, produced ethyl α-glucoside from AF in 80% ethanol. The findings indicate that α-glucosidases catalyze trans-addition. This is the first report of α-glucosidase-associated glucose formation from AF, possibly contributing to the salvage pathway of unutilized AF.

Crystal structure of bifunctional aldos-2-ulose dehydratase/isomerase from Phanerochaete chrysosporium with the reaction intermediate ascopyrone M.

The enzyme aldos-2-ulose dehydratase/isomerase (AUDH) participates in carbohydrate secondary metabolism, catalyzing the conversion of glucosone and 1,5-d-anhydrofructose to the secondary metabolites cortalcerone and microthecin, respectively. AUDH is a homo-dimeric enzyme with subunits of 900 amino acids. The subunit consists of a seven-bladed ß-propeller domain, two cupin folds and a C-terminal lectin domain. AUDH contains a structural Zn(2+) and Mg(2+) located in loop regions and two zinc ions at the bottom of two putative active-site clefts in the propeller and the cupin domain, respectively. Catalysis is dependent on these two zinc ions, as their specific removal led to loss of enzymatic activity. The structure of the Zn(2)(+)-depleted enzyme is very similar to that of native AUDH, and structural changes upon metal removal as the cause for the catalytic deficiencies can be excluded. The complex with the reaction intermediate ascopyrone M shows binding of this compound at two different sites, with direct coordination to Zn(2+) in the propeller domain and as second sphere ligand of the metal ion in the cupin domain. These observations suggest that the two reactions of AUDH might be catalyzed in two different active sites, about 60 Å apart. The dehydration reaction most likely follows an elimination mechanism, where Zn(2+) acts as a Lewis acid polarizing the C2 keto group of 1,5-d-anhydrofructose. Abstraction of the proton at the C3 carbon atom and protonation of the leaving group, the C4 hydroxyl moiety, could potentially be catalyzed by the side chain of the suitably positioned residue His155.

A simple and fast kinetic assay for phytases using phytic acid-protein complex as substrate.

Phytase (EC 3.1.3.-) hydrolyzes phytate (IP(6)) present in cereals and grains to release inorganic phosphate (P(i)), thereby making it bioavailable. The most commonly used method to assay phytase, developed nearly a century ago, measures the P(i) liberated from IP(6). This traditional endpoint assay is time-consuming and well known for its cumbersomeness in addition to requiring extra caution for handling the toxic regents used. This article reports a simple, fast, and nontoxic kinetic method adaptable for high throughput for assaying phytase using IP(6)-lysozyme as a substrate. The assay is based on the principle that IP(6) forms stable turbid complexes with positively charged lysozyme in a wide pH range, and hydrolysis of the IP(6) in the complex is accompanied by a decrease in turbidity monitored at 600 nm. The turbidity decrease correlates well to the released P(i) from IP(6). This kinetic method was found to be useful in assaying histidine acid phytases, including 3- and 6-phytases, a class representing all commercial phytases, and alkaline ß-propeller phytase from Bacillus sp. The influences of temperature, pH, phosphate, and other salts on the kinetic assay were examined. All salts, including NaCl, CaCl(2), and phosphate, showed a concentration-dependent interference.

Study on administration of 1,5-anhydro-D-fructose in C57BL/6J mice challenged with high-fat diet.

1,5-Anhydro-D-fructose (AF) is a mono-saccharide directly formed from starch and glycogen by the action of α-1,4-glucan lyase (EC 4.2.2.13). Our previous study has indicated that AF increases glucose tolerance and insulin secretion in NMRI mice after administration through a gastric gavage in a single dose at 150 mg per mouse. In this study, we used high-fat feeding of C57BL/6J mice to examine the influence of long-term administration of AF on glucose-stimulated insulin secretion in vivo and in vitro. We found that 8-weeks of high-fat feeding increased body weight, fasting blood glucose and insulin levels in C57BL/6J mice when compared to mice fed normal diet. Impaired glucose tolerance was also observed in mice receiving 8-weeks of high-fat diet. In contrast, AF (1.5 g/kg/day), administered through drinking water for 8-weeks, did not affect body weight or food and water intake in mice fed either the high-fat or normal diet. There was no difference in basal blood glucose or insulin levels between AF-treated and control group. Oral glucose tolerance test (OGTT) showed that AF did not affect glucose-stimulated insulin secretion in mice. In in vitro studies with isolated islets, AF did not influence glucose-stimulated insulin secretion in mice receiving either high-fat or normal diet. We therefore conclude that when given through drinking water for 8 weeks at 1.5 g/kg/day, AF has no effect on glucose-stimulated insulin secretion in C57BL/6J mice challenged with a high-fat diet.

1,5-anhydro-D-fructose and its derivatives: biosynthesis, preparation and potential medical applications.

1,5-Anhydro-D-fructose (AF) was first found in fungi and red algae. It is produced by the degradation of glycogen, starch and maltosaccharides with α-1,4-glucan lyase (EC 4.2.2.13). In vivo, AF is metabolized to 1,5-anhydro-D-glucitol (AG), ascopyrone P (APP), microthecin and other derivatives via the anhydrofructose pathway. The genes coding for the enzymes in this pathway have been cloned, enabling the large-scale production of AF and related products in a cell-free reactor. The possible applications of these products in medicine have been evaluated using both in vitro and in vivo systems. Thus AF is a useful anticariogenic agent as it inhibits the growth of the oral pathogen Streptococcus mutans, impairing the production of plaque-forming polysaccharides and lactic acid. AF also shows anti-inflammatory and anticancer effects. AG is used as a diabetic marker for glycemic control. AG also stimulates insulin secretion in insulinoma cell lines. in vivo, APP has been shown to lengthen the life span of cancer-afflicted mice. It interferes with tumor growth and metastasis by its cidal effects on fast multiplying cells. Microthecin inhibits the growth of the human pathogen Pseudomonas aeruginosa PAO1, particularly under anaerobic conditions. The pharmaceutical usefulness of the other AF metabolites 1,5-anhydro-D-mannitol,1-deoxymannojirimycin, haliclonol, 5-epipentenomycin I, bissetone, palythazine, isopalythazine, and clavulazine remains to be investigated. In this review AF and its metabolites as the bioactive natural products for their pharmaceutical potentials are discussed.

1,5-Anhydro-D-fructose: biocatalytic and chemical synthetic methods for the preparation, transformation and derivatization.

1,5-Anhydro-D-fructose (1,5AnFru) is a monoketosaccharide that can be prepared enzymatically from starch by alpha-1,4-glucan lyase or chemically from D-glucose or D-fructose in a few steps with high yields. The formed 1,5AnFru can be derivatized both enzymatically and chemically to interesting new carbohydrate derivatives, some with biological activities. For example dehydratases, isomerases and reductases can convert 1,5AnFru to enolones (as Ascopyrone P) and sugar alcohols with antimicrobial and antioxidant properties, while chemical modifications can give similar compounds as well as natural products like 1-deoxymannonojirimycin and Clavulazine. 1,5AnFru disaccharides (glycosyl 1-->4 1,5AnFru) have been prepared as well as glycosyl 1-->4 1,5-anhydro-D-tagatose.

[Photolumimescence character of novel phthalocyanine].

In the present paper, the authors study the photolumimescence spectra of the novel 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine casting film and vacuum-deposited film. Photolumimescence spectras of casting film on the quartz substrate were measured at 10, 77, 177 and 300 K, and the photolumimescence spectra of vacuum-deposited film with a thickness of about 200 nm on the silicon substrate was studied at room temperature (300 K). For 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine, the casting films all show fluorescence peaks at 942, 937, 942 and 942 nm and phosphorescence peaks at 1114, 1057, 1114 and 1114 nm in the photolumimescence spectra at 10, 77, 177 and 300 K, respectively. In the cases of 2,3-tetra-(2-isopropyl-5-methyl -benzoyl) hydrogen phthalocyanine, the peaks of excimers, which are related with the resistance ability of molecular aggregation, were found around 1673 nm as observed from photolumimescence spectra of the novel phthalocyanine casting films at 177 and 300 K. And the peak of excimers at 300 K is stronger than at 177 K also as can be seen from photolumimescence spectra of its casting films. With the increase in the temperature, the fluorescence peak was weakened and the peaks of excimers became stronger from the photoluminescence spectra of 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine casting films at 10, 77, 177 and 300 K. At the same time, the authors discussed the reason for coming into being 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine excimers as can be concluded from the structure of 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine molecules through the parameters of Chem 3D Ultra 9.0 MM2 calculation and simulated diagram of C4h isomer of 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine. The peaks of casting film and vacuum-deposited film of 2,3-tetra-(2-isopropyl-5-methyl -benzoyl) hydrogen phthalocyanine presented different maximum emission wavelength and full width at half maximum. The peak of 2,3-tetra-(2-isopropyl-5-methyl-benzoyl) hydrogen phthalocyanine vacuum-deposited films displays the maximum emission wavelengths around 1140 nm, while the maximum emission wavelengths of casting films show obvious differences compared with the vacuum-deposited films. The usual full width at half maximum is approximately 300 nm for casting film, which is in contrasts with that the full width at half maximum is about 100 nm for the vacuum-deposited film as can be seen from photolumimescence spectra of 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine casting film and photolumimescence spectra of 2,3-tetra-(2-isopropyl-5-methylbenzoyl) hydrogen phthalocyanine vacuum-deposited film.