Assessment of GH1, CAPN1 and CAST polymorphisms as markers of carcass and meat traits in Bos indicus and Bos taurus–Bos indicus cross beef cattle

The objective of this study was to investigate the effects of bovine GH1, CAPN1 and CAST gene polymorphisms on carcass and meat traits in Nellore and Nellore x Bos taurus beef cattle. Three hundred animals were genotyped for GH1/MspI (TC/G in intron 3), CAPN316 (AF_252504.2:g.5709C > G) and CAST/RsaI (AY_008267.1:g282C > G) and phenotyped for rib eye area, backfat thickness, intramuscular fat, shear force (SF), and myofibrillar fragmentation index (MFI). No significant associations were observed between the GH1/MspI and CAST/RsaI polymorphisms and phenotypes, although the relation between the CAST/RsaI genotypes and meat tenderness evaluated by MFI approached significant. The fact that the CAPN316 polymorphism did not show adequate segregation in Nellore cattle confirms the difficulty of using this marker in breeding programs of different Bos indicus breeds. However, the positive results of the association analysis obtained for Nellore x B. taurus crosses contributed to the validation of previous findings.     

Volatiles of the Balkan endemic Daucus guttatus ssp. zahariadii and cultivated and wild-growing D. carota – A comparison study

The hydrodistilled essential oil obtained from the aerial parts of Daucus guttatus Sibth. & Sm. ssp. zahariadii Heywood, an endemic plant species of the Balkan Peninsula, as well as solvent extracts and essential oils from different parts of Daucus carota L. have been analysed by GC and GC–MS and screened for antimicrobial activity against 12 bacterial and two fungal strains. The volatiles of the two plant taxa differed significantly in both their chemical identity and antimicrobial effect. The dominant constituent of D. guttatus oil was apiol (43.3%), which was absent from all samples of D. carota. The diethyl ether extract of D. carota inhibited the growth of the yeast Candida albicans while the oil of D. guttatus at 25 mg/ml had no effect on the growth of the fungal organisms tested. Additionally, the oil of D. guttatus showed prominent antibacterial activity against a pathogenic Corynebacterium pyogenes.     

In vitro and in vivo assessment of cytochrome P450-mediated herb–drug interaction of Ssang-hwa-tang

We have evaluated the herb–drug interaction potential of Ssang-hwa-tang (SHT) mediated by cytochrome P450 (CYP) inhibition/induction. Further, the effects of fermentation on the CYP-mediated herb–drug interaction potential were determined. SHT showed inhibitory activity toward CYP1A2, but not 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, and 3A4 in human liver microsomes. The results of the enzyme kinetic study suggested that the SHT-induced CYP1A2 inhibition is mixed reversible inhibition. The hepatic CYP expression and activity in rats treated with SHT were examined. The expression/activity of CYP2E1 increased as a result of SHT extract treatment (P < 0.005 or P < 0.001, respectively), which raises the possibility that SHT may increase the toxicity of environmental toxicants through the elevation of CYP2E1-mediated metabolic activation. SHT fermentation using Lactobacillus fermentum or Lactobacillus gasseri resulted in attenuation of the SHT-induced CYP1A2 inhibition, but not CYP2E1 induction, suggesting that changes in the chemical composition of SHT through fermentation can affect the inhibition of CYP1A2 activity.     

Bioassay-guided screening and isolation of α-glucosidase and tyrosinase inhibitors from leaves of Morus alba

In this study, bioassay-guided fractionation of extracts from the leaves of Morus alba L. led to the isolation of 15 bioactive constituents with α-glucosidase and tyrosinase inhibitory activities, among which prenylated stilbenes were proved to be a new group of α-glucosidase inhibitors apart from iminosugars derived from Morus alba. Their structures were identified on the basis of extensive spectroscopic analysis and chemical evidence, as well as comparing with data from the literature. Among them, compounds (2R)/(2S)-Euchrenone a₇ (6a/6b), Chalcomoracin (7), Moracin C (8), Moracin D (9) and Moracin N (10) exhibited a significant degree of α-glucosidase inhibitory activity with IC₅₀ of 6.28, 2.59, 4.04, 2.54 and 2.76 μM, respectively, while (2R)/(2S)-Euchrenone a₇ (6a/6b), Moracin N (10), Quercetin (13), Norartocarpetin (14), the interconvertible epimeric mixture of (2R)/(2S)-7-methoxyl-8-ethyl-2′,4′-dihydroxylflavane-2″-O-β-d-glucopyranoside (1a/1b) and the interconvertible enantiomers of (2R)/(2S)-7-methoxyl-8-hydroxyethyl-2′,4′-dihydroxylflavane (5a/5b) displayed a potent tyrosinase inhibitory effect with IC₅₀ of 0.260, 0.924, 0.523, 0.0824, 0.616 and 0.528 μM, respectively. Especially, (2R)-7-methoxyl-8-ethyl-2′,4′-dihydroxylflavane-2″-O-β-d-glucopyranoside (1a), (2S)-7-methoxyl-8-ethyl-2′,4′-dihydroxylflavane-2″-O-β-d-glucopyranoside (1b), (2S)-8-hydroxyethyl-7,4′-dimethoxylflavane-2′-O-β-d-glucopyranoside (2), (2R)-7-methoxyl-8-hydroxyethyl-2′,4′-dihydroxylflavane (5a) and (2S)-7-methoxyl-8-hydroxyethyl-2′,4′-dihydroxylflavane (5b) were identified as new compounds.     

Impact of the co-culture of Saccharomyces cerevisiae–Oenococcus oeni on malolactic fermentation and partial characterization of a yeast-derived inhibitory peptidic fraction

The present study was aimed to evaluate the impact of the co-culture on the output of malolactic fermentation and to further investigate the reasons of the antagonism exerted by yeasts towards bacteria during sequential cultures. The Saccharomyces cerevisiae D strain/Oenococcus oeni X strain combination was tested by applying both sequential culture and co-culture strategies. This pair was chosen amongst others because the malolactic fermentation was particularly difficult to realize during the sequential culture. During this traditional procedure, malolactic fermentation started when alcoholic fermentation was achieved. For the co-culture, both fermentations were conducted together by inoculating yeasts and bacteria into a membrane bioreactor at the same time. Results obtained during the sequential culture and compared to a bacterial control medium, showed that the inhibition exerted by S. cerevisiae D strain in term of decrease of the malic acid consumption rate was mainly due to ethanol (75%) and to a peptidic fraction (25%) having an MW between 5 and 10 kDa. 0.4 g l⁻¹ of l-malic acid was consumed in this case while 3.7 g l⁻¹ was consumed when the co-culture was applied. In addition, there was no risk of increased volatile acidity during the co-culture. Therefore, the co-culture strategy was considered effective for malolactic fermentation with the yeast/bacteria pair studied.     

Substrate specificities and mechanism of action of multiple α-galactosidases from Streptomyces griseoloalbus

α-Galactosidase or melibiase is a versatile enzyme with many potential biotechnological and industrial applications. The substrate specificities of three α-galactosidases – α-Gal I, α-Gal II, and α-Gal III – from Streptomyces griseoloalbus were studied using different galactose-containing natural substrates like melibiose, raffinose and stachyose. The kinetic parameters K_m and V_max were determined from the Lineweaver–Burk plot. α-Gal I showed highest affinity towards melibiose where as α-Gal II and α-Gal III showed highest affinity towards stachyose. The ¹H NMR studies showed that all the three α-galactosidases had a retaining mechanism of hydrolysis.     

Fractionation of lipids and purification of γ-linolenic acid (GLA) from Spirulinaplatensis

The microalgae, Spirulinaplatensis, is an excellent source of γ-linolenic acid (GLA), an essential polyunsaturated fatty acid and a potent nutraceutical. The fatty acid composition of S.platensis ARM 740 was determined after transmethylation by gas chromatography (GC). Lipid fractionation was achieved on silica gel column chromatography and preparative TLC. Neutral lipids, glycolipids and phospholipids accounted for 77.0%, 15.6% and 7.4%, respectively, of the total lipid fraction. S.platensis ARM 740 was found to contain 94% of the total GLA in the glycolipid fraction. Attempts were made to purify GLA methyl ester by using urea to form inclusion complexes with the saturated and the less unsaturated FAMEs (fatty acid methyl esters), which enhanced the purity of GLA methyl ester to 84%. A further approach to isolate GLA methyl ester with higher purity involved the use of argentated silica gel chromatography. An initial PUFA concentration step frequently adopted by most researchers to increase GLA purity was not necessary in the isolation of GLA from S.platensis. A GLA methyl ester with a purity of >96% and a recovery of 66% was obtained. Purity of the isolated GLA methyl ester was confirmed by GC and IR analysis with respect to authentic standard.     

Enzymatic properties and transglycosylation of α-galactosidase from Penicillium oxalicum SO

Penicillium oxalicum SO α-galactosidase demonstrated weak hydrolysing activity but a high rate of transglycosylation in the reaction with melibiose, where the major product was 6-α-galactosyl melibiose. The transfer ratio was 83.6% and was maintained over a long reaction time of 80 h. The molecular weight was estimated to be 124,000 by SDS–PAGE. The optimal pH was ∼3 and a stable pH, with a range of 2.4–9.5, was found. The optimal temperature was ∼60 °C and the activity was stable below 60 °C. With respect to acceptor specificity, mono-alcohols, sugar alcohols and sugars were poor acceptors, but the di-alcohol ethylene glycol and the tri-alcohol glycerin were good acceptors. The percentage of transglycosylation to glycerin increased up to 41.7%, as that to melibiose decreased, with the initial glycerin concentration of 40%. The production of α-d-galactosylglycerol was 293 mg for each gram of melibiose used by the enzymatic reaction.     

Acidification,crushing and thermal treatments can influence the profile and stability of folate poly-γ-glutamates in broccoli (Brassica oleracea L. var. italica)

The influence of different treatments, i.e., crushing, high temperature short time (90 °C/4 min) (HTST) and low temperature long time (60 °C/40 min) (LTLT) blanching, acidification (pH 4.3), and sequences of these treatments on the folate poly-γ-glutamate profile and stability were investigated. In this study, broccoli was used as a case study. Regarding the folate poly-γ-glutamate profile, endogenous folate poly-γ-glutamates in broccoli florets were found predominantly as hepta- and hexa-γ-glutamates. Crushing raw broccoli, acidification and LTLT blanching enhanced folate deconjugation resulting in monoglutamate, di- and tri-γ-glutamates. Compared to other treatments, HTST blanching preformed prior to crushing resulted in the highest concentration of long chain poly-γ-glutamates. Regarding folate poly-γ-glutamates stability, acidification combined with LTLT blanching decreased folate stability whereas HTST blanching combined with different sequences of blanching and crushing did not affect folate poly-γ-glutamates stability. It was concluded that crushing (prior to heating), acidification and blanching could be strategically applied to increase the folate monoglutamate content of broccoli.     

Occurrence of extended-spectrum β-lactamases among isolates of Salmonella enterica subsp. enterica from food-producing animals and food products,in Portugal

A total of 1120 Salmonella spp. isolates, recovered from poultry, swine and food products of animal origin (bovine, swine and poultry) over the period of 2009–2011, were investigated in order to determine their serotype, susceptibility to a panel of eleven antimicrobials (A, ampicillin; Ct, cefotaxime; Cp, ciprofloxacin; Tm, trimethoprim; Su, sulfamethoxazole; C, chloramphenicol; S, streptomycin; G, gentamicin; T, tetracycline; NA, nalidixic acid; Fl, florfenicol), and the presence of resistance determinants of extended-spectrum cephalosporins. Overall, Salmonella Enteritidis was the most common serotype in all three animal species. In 618 isolates of poultry, 32.8% comprised S. Enteritidis, 18.3% Salmonella Havana and 16.5% Salmonella Mbandaka; in 101 isolates of pigs, 21.8% comprised Salmonella Rissen and Salmonella Typhimurium, 10.9% Salmonella Derby and Salmonella London. Salmonella I 4,[5],12:i:- was the most common serotype recovered from pork and beef food products comprising 32.6% and 30% of isolates respectively, followed by S. Rissen (26% and 24%) and S. Typhimurium (18.2% and 19%), respectively. In poultry products, S. Enteritidis was the most frequent serotype (62.7%), followed by S. Mbandaka (10.2%) and S. Derby (8.5%). Susceptibility profiles differed according to the origin of the isolates. Five multidrug resistant isolates (0.45%) were further characterized as extended-spectrum β-lactamase (ESBL) producers. Polymerase chain reaction and sequencing of the amplicons confirmed the presence of bla_CTX-M-1 (n = 2), bla_CTX-M-14 (n = 1), bla_CTX-M-15 (n = 1) and bla_CTX-M-32 (n = 1); bla_SHV-12 and bla_TEM-1 genes were also detected in two isolates of S. I 4,[5],12:i:-. Four isolates, two S. Havana and two S. I 4,[5],12:i:-, carried class 1 integrons and in three, two S. I 4,[5],12:i:- and one S. Havana, ISEcp1 was identified associated to bla_CTX-M-1, bla_CTX-M-32 and bla_CTX-M-14 genes. Additionally, in one S. I 4,[5],12:i:- isolate, orf477 was identified linked to bla_CTX-M-32. No plasmid mediated quinolone resistance-encoding genes were detected. Here, we report for the first time the presence of bla_CTX-M genes in Salmonella enterica subsp enterica isolates recovered from poultry and food products of swine origin, in Portugal.     

Comparing the in vitro effects of MGO™ Manuka honey and tea tree oil on ocular Demodex viability

Purpose

To compare the in vitro antiparasitic effects of MGO^? Manuka honey and tea tree oil against ocular Demodex.

Antioxidant and antiacetylcholinesterase activities of essential oils from Cymbopogon schoenanthus L. Spreng. Determination of chemical composition by GC–mass spectrometry and C NMR

Cymbopogon schoenanthus L. Spreng, is an aromatic herb consumed in salads and used to prepare traditional meat recipes in Tunisia. The chemical composition, antioxidant activities and acetylcholinesterase inhibitory properties of the essential oils from fresh leaves, dried leaves and roots collected from three different locations in southern Tunisia, were evaluated. Essential oils were analysed by GC–mass spectrometry and ¹³C NMR. The major components were limonene (10.5–27.3%), β-phellandrene (8.2–16.3%), δ-terpinene (4.3–21.2%) and α-terpineol (6.8–11.0%). Antioxidant activity was measured by DPPH assay. The results ranged from 36.0% to 73.8% (2 μl of essential oil per mL of test solution).     

Volatile profiling of Ficus carica varieties by HS-SPME and GC–IT-MS

Aroma is one of the essential parameters for the evaluation of fruit quality, with volatile components being determinant for this characteristic. During this work, the volatile profile of fresh fruits (pulp and peel) and leaves of Portuguese Ficus carica L. white (“Pingo de Mel” and “Branca Tradicional”) and dark (“Borrasota Tradicional”, “Verbera Preta” and “Preta Tradicional”) varieties were characterised by HS-SPME/GC–IT-MS.     

Solid-state fermentation of apple pomace using Phanerocheate chrysosporium – Liberation and extraction of phenolic antioxidants

Apple pomace is a by-product from the apple processing industry and can be used for the production of value-added phenolic compounds. A study was carried out to understand the changes and liberation of phenolic compounds and improvement in antioxidant activity during solid-state fermentation of apple pomace using Phanerocheate chrysosporium. The solid-state fermentation of apple pomace using P. chrysosporium mobilised the polyphenolic compounds and improved the nutraceutical properties. The polyphenol content in acetone extract increased and the results were statistically significant (P < 0.05) from 4.6 to 16.12 mg GAE/g dry weight during solid-state fermentation. The effect of various solvents, temperature, time and detergents were also investigated for the extraction of polyphenolics by ultrasonication and microwave-assisted extraction methods. The polyphenol content of the extracts was found to be in the range of 5.78–16.12 mg GAE/g DW of samples, depending on the solvent, extraction time and temperature. Antioxidant activities of polyphenol extracts were tested using the 2,2-diphenyl-1-picryhydrazyl (DPPH) radical methods, where the IC₅₀ ranged from 12.24 to 23.42 μg DW sample, depending on the extraction conditions and the antioxidant activities correlated well with the polyphenol concentrations.     

Effect of Co γ-irradiation on postharvest quality and selected enzyme activities of Pleurotus nebrodensis

Freshly harvested Pleurotus nebrodensis fruit bodies were exposed to four different doses (0.8-2.0 kGy) of ⁶⁰Co γ-irradiation and various physiological changes associated with postharvest deterioration, as well as the activities of selected enzymes (proteinase, polyphenoloxidase, superoxide dismutase, catalase) thought to play a role in the process of deterioration, were monitored during 22 days of subsequent storage at 4 °C and 65-70% relative humidity. An irradiation dose of 1.2 kGy significantly delayed (by 6-9 days) the onset of fruit body softening, splitting and browning compared with non-irradiated controls and test samples subjected to lower or higher irradiation doses. Irradiation with 1.2 and 1.6 kGy also had a positive effect on other indicators of mushroom tissue senescence, resulting in smaller decreases in soluble protein levels and more protracted increases in proteinase activity. Peak levels of polyphenoloxidase activity, widely recognized as causing postharvest browning of mushroom tissue, were also significantly lower (P < 0.05) in fruit bodies exposed to 1.2 kGy compared with non-irradiated controls. Our data increase our understanding of the effects of γ-irradiation on the biochemical changes associated with postharvest deterioration in P. nebrodensis, and improve the prospects of more targeted strategies for extending the shelf life of both this and other mushrooms.     

Enzymatic synthesis of fructooligosaccharides by inulinases from Aspergillus niger and Kluyveromyces marxianus NRRL Y-7571 in aqueous–organic medium

This work is focused on the synthesis of the fructooligosaccharides (FOS) from sucrose and inulin, using free, immobilized and pre-treated immobilized inulinase from Kluyveromyces marxianus NRRL Y 7571 and Aspergillus niger in an aqueous–organic system. Initially, the influence of pre-treatment using four different gases, propane, n-butane, CO₂ and liquefied petroleum gas (LPG), was investigated towards FOS production and best results were found when both enzymes were previously treated with LPG. The best reaction yields were obtained when the immobilized enzymes were treated with LPG. Considering FOS synthesis using the enzyme from A. niger, yields of 26.62% of GF2 (kestose), 30.62% of GF3 (nystose) and 8.47% of GF4 (fructosyl nystose) were achieved using sucrose as substrate. Using inulinases from K. marxianus NRRL Y 7571, 11.89% of GF2 and 20.83% of GF3 were obtained, using inulin as substrate. However, promising results were achieved using the free form of inulinase from A. niger (77.19% of GF2; 14.03% of GF3 and 0.07% of GF4) using inulin as substrate.     

HPLC–MS identification and quantification of flavonol glycosides in 28 wild and cultivated berry species

Berries and red fruits are rich dietary sources of polyphenols with reported health benefits. More than 50 different flavonols (glycosides of quercetin, myricetin, kaempferol, isorhamnetin, syringetin and laricitrin) have been detected and quantified with HPLC–MSⁿ in fruits of blueberry, bilberry, cranberry, lingonberry, eastern shadbush, Japanese wineberry, black mulberry, chokeberry, red, black and white currants, jostaberry, red and white gooseberry, hardy kiwifruit, goji berry, rowan, dog rose, Chinese and midland hawthorn, wild and cultivated species of blackberry, raspberry, strawberry and elderberry. The phenolic constituents and contents varied considerably among the analyzed berry species. Elderberry contained the highest amount of total flavonols (450–568 mg kg⁻¹ FW), followed by berry species, containing more than 200 mg kg⁻¹ FW of total: chokeberry (267 mg kg⁻¹), eastern shadbush (261 mg kg⁻¹), wild grown blackberry (260 mg kg⁻¹), rowanberry (232 mg kg⁻¹), american cranberry (213 mg kg⁻¹) and blackcurrants (204 mg kg⁻¹). Strawberry (10.5 mg kg⁻¹) and white currants (4.5 mg kg⁻¹) contained the lowest amount of total flavonols. Quercetins represent the highest percentage (46–100%) among flavonols in most analyzed berries. In wild strawberry and gooseberry the prevailing flavonols belong to the group of isorhamnetins (50–62%) and kaempferols, which represent the major part of flavonols in currants (49–66%). Myricetin glycosides could only be detected in chokeberry, rowanberry and species from the Grossulariaceae, and Adoxaceae family and Vaccinium genus. Wild strawberry and blackberry contained from 3- to 5-fold higher total flavonols than the cultivated one.     

Carbohydrate and protein sources influence the induction of α- and β-galactosidases in Lactobacillus reuteri

The induction of α- and β-galactosidases in six strains of Lactobacillus reuteri (L. reuteri) by six carbohydrate sources and four protein sources was studied. L. reuteri grown on raffinose had the highest α-galactosidase activity (10.55 Gal U/ml), while lactose exhibited the highest β-galactosidase activity (43.82 Gal U/ml) when compared to other carbohydrate sources. L. reuteri grown on yeast extract exhibited the highest α- and β-galactosidases activity (15.27 and 12.88 Gal U/ml, respectively) when compared to other protein sources. MF14C and SD2112 grown on raffinose had the highest α-galactosidase activity (14.75 and 14.18 Gal U/ml, respectively) followed by CF2-7F (13.38 Gal U/ml). CF2-7F grown on lactose had the highest β-galactosidase activity (82.01 Gal U/ml). SD2112, MM2-3 and CF2-7F grown on yeast extract (20.96, 19.67, 19.67 Gal U/ml, respectively) showed the highest α-galactosidase activity. MM2-3 and CF2-7F grown on yeast extract showed the highest β-galactosidase activity (18.1 and 17.59 Gal U/ml, respectively). Raffinose and lactose were the best carbohydrate sources to produce α- and β-galactosidases, respectively. Yeast extract was the best protein source to produce both enzymes and CF2-7F strain was the best producing strain on all tested conditions.     

Effect of lyoprotectants on β-glucosidase activity and viability of Bifidobacterium infantis after freeze-drying and storage in milk and low pH juices

The benefit of disaccharide protectants for maintaining viability and β-glucosidase activity of Bifidobacterium infantis UV16PR during freeze-drying and storage in different food matrices was investigated. Protectants used were cellobiose, lactose, sucrose and trehalose.     

Characterization of the components present in the active fractions of health gingers (Curcuma xanthorrhiza and Zingiber zerumbet) by HPLC–DAD–ESIMS

Curcuma xanthorrhiza and Zingiber zerumbet are two of the most commonly used ingredients in Indo-Malaysian traditional medicines, health supplements and tonics. Recently, a number of products derived from the aqueous extracts of these species have appeared in the market in the form of spray-dried powder packed in sachet or bottle. On-line high performance liquid chromatography, coupled with diode array detection and electrospray ion trap tandem mass spectroscopy (HPLC–DAD–ESI–MSⁿ), was used to analyze the components in the antioxidant-active fractions from the rhizomes of these species. Three components were identified from C. xanthorrhiza, including bisdemethoxycurcumin (1), demethoxycurcumin (2) and curcumin (3). The active fraction from Z. zerumbet consisted of five components, including kaempferol 3-O-rhamnoside (4), compound 5 [kaempferol 3-O-(2″-O-acetyl)rhamnoside (5a) or kaempferol 3-O-(3″-O-acetyl)rhamnoside (5b)], kaempferol 3-O-(4″-O-acetyl)rhamnoside (6), kaempferol 3-O-(3″,4″-O-diacetyl)rhamnoside (7) and kaempferol 3-O-(2″,4″-O-diacetyl)rhamnoside (8). To confirm their identities, the components from Z. zerumbet were isolated conventionally and were analyzed by spectroscopic techniques as well as by comparison with literature data.