M. Shuhaimi

Purification of serine protease from mango (Mangifera Indica Cv. Chokanan) peel using an alcohol/salt aqueous two phase system

An alcohol/salt-based aqueous two-phase (ATPS) system, as a novel method of purification, was employed to purify serine proteases from mango (Mangifera Indica Cv. Chokanan) peel. The effectiveness of different parameters, such as type and concentration of alcohol (1-propanol, 2-propanol, and ethanol), type of salt (sodium citrate, potassium phosphate, and ammonium sulphate), pH, and NaCl, on the purification and selective separation of serine protease was investigated. Desirable conditions of partition coefficient (K), selectivity (S), purification factor (P), and yield (Y%) of serine protease, using ATPS, were determined. The highest partition coefficient (64.5) and selectivity (343.2) for serine protease purification value were achieved in an ATPS of 16% (w/w) 2-propanaol, 19% (w/w) potassium phosphate, and 5% (w/v) NaCl at pH 7.5. It was demonstrated that serine protease could be recovered with a yield of 96.7% and a purification factor of 11.6.

Selected microbial groups and short-chain fatty acids profile in a simulated chicken cecum supplemented with two strains of Lactobacillus

Among the bacterial fermentation end products in the chicken cecum, butyrate is of particular importance because of its nutritional properties for the epithelial cell and pathogen inhibitory effects in the gut. An in vitro experiment, operated with batch bioreactor, was conducted to quantify butyric-producing bacteria in a simulated broiler cecum supplemented with Lactobacillus salivarius ssp. salicinius JCM 1230 and Lactobacillus agilis JCM 1048 during 24 h of incubation. Selected bacterial species were determined by real-time PCR and short-chain fatty acids and lactate concentrations were monitored. The results showed that after 24 h of incubation, Lactobacillus supplementation significantly increased the number of lactobacilli, bifidobacteria and Faecalibacterium prausnitzii in medium containing cecal content and lactobacilli supplementation (Cc + L) compared with the control (Cc). Addition of lactobacilli did not alter Escherichia coli and Clostridium butyricum, whereas it significantly (P < 0.05) reduced Salmonella in treatment Cc + L compared with the Cc treatment. Propionate and butyrate formation were significantly (P < 0.05) increased in treatment Cc + L as compared with the Cc treatment. Lactate was only detected in treatment containing 2 Lactobacillus strains. After 24 h of incubation, acetate concentration significantly (P < 0.05) decreased in all treatments. It was suggested that lactate produced by Lactobacillus in the cecal content improved the growth of butyric producers such as F. prausnitzii, which significantly increased butyrate accumulation. Additionally, the results showed that butyrate and propionate inhibited Salmonella without influencing the E. coli profile.

Utilisation of enterobacterial repetitive intergenic consensus (ERIC) sequence-based PCR to fingerprint the genomes of Bifidobacterium isolates and other probiotic bacteria

Enterobacterial repetitive intergenic consensus based on PCR (ERIC-PCR) was used to generate DNA fingerprints for bifidobacteria and other probiotic bacteria. Two primers (ERIC 1R and ERIC 2) used in ERIC-PCR revealed that all of the probiotic bacteria tested possess enterobacterial repetitive intergenic consensus sequences with the PCR products ranging from 250 bp to 5000 bp. The bacterial strains can be differentiated by comparing fingerprint patterns. The dendrogram of the fingerprints revealed that most of the bifidobacterial wild type strains fell into one cluster at similarity level of approximately 79%.

In vitro fermentation of broiler cecal content: the role of lactobacilli and pH value on the composition of microbiota and end products fermentation

Aim:  To assess the probiotic effects of Lactobacillus agilis JCM 1048 and L. salivarius ssp. salicinius JCM 1230 and the pH on the cecal microflora of chicken and metabolic end products.

Growth of Bifidobacterium longum BB536 in medida (fermented cereal porridge) and their survival during refrigerated storage

Aims:  To develop medida, a Sudanese fermented thin porridge as a probiotic dietary adjunct with high total solids.

Gastrointestinal tract morphological alteration by unpleasant physical treatment and modulating role of Lactobacillus in broilers

1. An experiment was conducted to determine the effects of supposedly unpleasant physical treatment on broiler performance, small intestinal development and ameliorating role of probiotics. 2. The following treatments were applied from day one: (1) chicks exposed to normal human contact fed basal diet (control); (2) chicks were exposed to unpleasant physical treatment and fed basal diet (UPT-BD); and (3) chicks were exposed to unpleasant physical treatment and fed basal diet supplemented with Lactobacillus (UPT-BDL). Chicks were exposed to UPT from days 1 to 21. Different segments of gastrointestinal tract were sampled at 14, 28, 35 and 42 d of age. 3. Broilers of UPT-BD had lower feed consumption compared with control group at 7 d of age. Overall, UPT-BDL birds showed higher body weight gain (BWG) and better feed conversion ratio (FCR) over the course of the experiment. 4. Birds of UPT-BD had lower concentrations of lactic, propionic and butyric acids in the caecum as compared with other groups at 14 d of age. Acetic acid concentration was profoundly decreased in both UPT groups compared to the control. 5. Duodenal villus height of UPT-BD broilers showed a slight reduction compared to the control and UPT-BDL birds at 14 d of age. Afterwards until day 42, UPT-BDL birds showed the highest villus height among treatments in different parts of the small intestine. 6. The results suggested that, even though UPT did not have significant inhibitory effects on the development of the small intestine and broiler performance, it negatively affected bacterial metabolic end products in the caecum, which could be ameliorated by the addition of Lactobacillus.

Efflux genes and active efflux activity detection in Malaysian clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA).

Efflux‐mediated resistance has been recognized as an important contributor of antibiotic resistance in bacteria, especially in methicillin‐resistant Staphylococcus aureus (MRSA) isolates. This study was carried out to detect and analyze efflux genes (norA and mdeA) and active efflux activity in a collection of Malaysian MRSA and methicillin‐sensitive S. aureus (MSSA) clinical isolates. Nineteen isolates including three ATCC S. aureus reference strains were subjected to PCR detection and DNA sequence analysis for norA and mdeA and active efflux detection using modified minimum inhibitory concentration (MIC) assay. From the 19 isolates, 18 isolates harboured the mdeA gene while 16 isolates contained norA gene. DNA sequence analysis reveals 98–100% correlation between the PCR product and the published DNA sequences in GenBank. In addition, 16 isolates exhibited active efflux activity using the ethidium bromide (EtBr)‐reserpine combination MIC assay. To our knowledge, this is the first report on the detection of efflux genes and active efflux activity amongst Malaysian clinical isolates of MRSA/MSSA. Detection of active efflux activity may explain the previous report on efflux‐mediated drug resistance profile amongst the local clinical isolates. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Influence of pH and impeller tip speed on the cultivation ofBifidobacterium pseudocatenulatum G4 in a milk-based medium

Biomass production ofBifidobacterium pseudocatenulatum G4 in a milk-based medium was carried out in a 2- and 10-L stirred tank fermenters. The effects of impeller tip speed (0.28, 0.56, and 0.83 m/s) and pH control (6.0, 6.5, and 7.0) on the biomass production were investigated. The growth performance in the 2-L fermenter was significantly improved when the impeller tip speed was held constant at 0.56 m/s and the pH was controlled at 6.5. These conditions yielded a maximum biomass of 1.687×109 cfu/mL, a maximum specific growth rate of 0.504 h−1, a biomass productivity of 9.240×107 cfu/mL·h, and a biomass yield of 9.791×1010 cfu/g lactose. The consumption of milk lactose resulted in the accumulation of 7.353 g/L acetic acid and 6.515 g/L lactic acid, with an acetic:lactic ratio of 1.129. Scale-up of the fermentation process to a 10-L fermenter based on a constant impeller tip speed of 0.56 m/s yielded reproducible results with respect to biomass production and cell viability.

Cloning and sequence analysis of bile salt hydrolase (bsh) gene from Bifidobacterium longum

A pair of PCR primers for the rapid detection of bile salt hydrolase (bsh) gene from Bifidobacterium longum BB536 has been synthesised and have revealed the bsh gene of approx 970 bp in Bifidobacterium longum BB 536 but not in other species of bacteria tested. The bsh gene was cloned and sequenced showing a high similarity to bsh gene previously published. The resulting nucleotide sequence encodes a predicted protein of 317 amino acids, Mw = 35 kDa.

Authentication of butter from lard adulteration using high-resolution of nuclear magnetic resonance spectroscopy and high-performance liquid chromatography

ABSTRACT Food authentication is an interesting issue for all parties in the food industry, including the fats and oils industry. Some unethical players try to blend high-quality foods, such as butter, with lower ones like lard, therefore, the analytical methods capable of analyzing the adulteration practices must be developed. This study used proton nuclear magnetic resonance spectroscopy in combination with high-performance liquid chromatography for the authentication of butter from lard adulteration. The identification of triacylglycerol composition of lard as a chemical marker for halal authentication is analyzed using high-performance liquid chromatography and high resolution nuclear magnetic resonance spectroscopy. The suitability of proton nuclear magnetic resonance provides a high-performance approach for determination butter adulterated with lard in their entirety of all proton bearing components. Peaks in the region of 2.60–2.84 ppm show special characteristics only present in lard. Only lard has its own unique characteristics which only polyunsaturated fatty acids would give signals 7 at δ 2.63, that corresponded to the chemical shift of the double-allylic methylene protons. In the same way, the intensity of signal at 2.63 ppm, due to methylenic protons in a position α to two double bonds, that is to say, due to the linoleic group. Furthermore, we also correlate some signals between 1H and 13C-NMR spectra for the confirmation of signals.