Aran H-Kittikun

Prevalence of Vibrio cholerae and Salmonella in a major shrimp production area in Thailand

In 1992 and 1993, a 7 months study carried out in a major shrimp-producing area in Southern Thailand to study the prevalence of Vibrio cholerae and Salmonella. A total of 158 samples were examined including water, sediment, shrimp, pelleted feed, shrimp gut, and chicken manure. Salmonella was not recovered from any sample type studied. V. cholerae O1 was isolated from 2 (2%) and V. cholerae non-O1 was isolated from 35 (33%) of 107 samples examined. The occurrence of V. cholerae was not significantly influenced by water salinity, temperature, dissolved oxygen or pH. There was no correlation between fecal coliform counts and the prevalence of V. cholerae. The results indicate that V. cholerae non-O1 is ubiquitous in aquatic environments where shrimp culture is practised under a variety of environmental conditions. The public health significance of non-O1 V. cholerae in shrimp culture remains to be determined. V. cholerae O1 and Salmonella do not appear to constitute a hygienic problem even if chicken manure was used as fertilizer.

Screening for new hydroxynitrilases from plants

We established a simple HPLC method to determine the activity and stereochemistry of the chiral mandelonitrile synthesized from benzaldehyde and cyanide, and applied it to screen for hydroxynitrile lyase (HNL) activity of plant origin. A total of 163 species of plants among 74 families were examined for (R)- and (S)-HNL activities using the method. We discovered that homogenate of leaves of Baliospermum montanum shows (S)-HNL activity, while leaves and seeds from Passiflora edulis, and seeds from Eriobotrya japonica, Chaenomles sinensis, Sorbus aucuparia, Prunus mume, and Prunus persica show (R)-HNL activity. Partially purified (R)-HNLs from Passiflora edulis and Eriobotrya japonica acted not only on benzaldehyde but also on aliphatic ketone. The enantiomeric excess of (R)-methylpropylketone cyanohydrin synthesized from 2-pentanone using homogenate from leaves of Passiflora edulis was 87.0%, and that of (R)-mandelonitrile synthesized by homogenate from seeds of Eriobotrya japonica was 85.0%.

Hydroxynitrile lyase from Passiflora edulis: Purification, characteristics and application in asymmetric synthesis of (R)-mandelonitrile

A hydroxynitrile lyase from leaves of Passiflora edulis (PeHNL) was purified and characterized for the first time. The enzyme is a monomer of 15kDa and 18kDa by SDS-PAGE, and gel filtration, respectively. Asymmetric synthesis of (R)-mandelonitrile from benzaldehyde and acetone cyanohydrin in a biphasic system employing the PeHNL from rinds of P. edulis was carried out. Several parameters influenced the enantiomeric purity of the product and initial velocity of the reaction. Both pH and temperature were important parameters controlling the enantiomeric purity of the product. The optimum pH and temperature were pH 4 and 10°C, respectively. At the optimum pH and temperature, the spontaneous non-enzymatic reaction yielding the racemic mandelonitrile was almost completely suppressed. The PeHNL was stable (more than 80% residual activity after incubation for 12h) in the system of methyl-t-butyl ether (MTBE), dibutyl ether (DBE), hexane (HEX), and diisopropyl ether (DIPE) while diethyl ether (DEE) and ethyl acetate (EA) were not suitable solvents. The initial velocity was markedly affected by the type of organic solvent in the biphasic system, while high enantiomeric purity was obtained when organic solvents having logP lower than 3.5 were used. The highest initial velocity of reaction and enantiomeric purity of (R)-mandelonitrile were obtained in the biphasic system of DBE with the aqueous phase content of 30% (v/v). The optimum substrate concentrations were 250mM for benzaldehyde and 900mM for acetone cyanohydrin, and the optimum enzyme concentration was 26.7units/ml. The highest enantiomeric purity of (R)-mandelonitrile was successfully obtained with conversion and enantiomeric excess of 31.6% and 98.6%, respectively. The enzyme showed considerable reusability in batch reaction with high enantiomeric purity of product. Herein, we reported the characteristics of a unique (R)-PeHNL from leaves of P. edulis. The PeHNL from rinds had been isolated for the first time and the enzyme showed great ability in transcyanation of (R)-mandelonitrile with high e.e. in DBE as the co-organic solvent in a biphasic system.

Lipids from cephalothorax and hepatopancreas of Pacific white shrimp (Litopenaeus vannamei): Compositions and deterioration as affected by iced storage

Lipids from cephalothorax and hepatopancreas of Pacific white shrimp (Litopenaeus vannamei) stored in ice for up to 6 days were extracted and characterised. The extraction yields of lipids from hepatopancreas (10.65-12.64%) were higher than those from cephalothorax (2.59-2.88%). However, no changes in the extraction yield were observed during the storage (p>0.05). The carotenoid contents of lipids from cephalothorax and hepatopancreas slightly increased within the first 2 and 4 days of iced storage (p<0.05), respectively, but decreased thereafter (p<0.05). With increasing storage time, a progressive formation of hydroperoxide was found as evidenced by the increase in the absorbance band at 3600-3200 cm(-1) in Fourier transform infrared (FTIR) spectra, and increased peroxide values (PVs) (p<0.05). The increases in thiobarbituric acid reactive substances (TBARS), p-anisidine value (AnV) and free fatty acid (FFA) content of lipids were noticeable when iced storage time increased (p<0.05). Those changes indicated that lipid oxidation and hydrolysis occurred in both samples. Phospholipids (PL) were the major components in lipids from cephalothorax (82.51% of total lipids). Nevertheless, lipids from hepatopancreas contained triglyceride (TG) and PL as the dominant components (45.35% and 38.03% of total lipids, respectively). A decrease in the TG content with a concomitant increase in free fatty acid was observed at the end of storage (day 6) (p<0.05). Decreases in unsaturated fatty acids, especially eicosapentaenoic acid (EPA; C20:5(n-3)) and docosahexaenoic acid (DHA; C22:6(n-3)) were noticeable at day 6 of storage (p<0.05). Thus, the extended storage time resulted in the enhanced deterioration of extracted lipids.

Effect of lauric arginate, nisin Z, and a combination against several food-related bacteria.

The effects of lauric arginate (LAE) and nisin Z, alone or in combination, on cell damage were investigated against Escherichia coli O157:H7, Listeria monocytogenes and Brochothrix thermosphacta, by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) observations, efflux of potassium and phosphate ions, and growth inhibition. A combination of LAE with nisin Z caused severe and dramatic changes in the cytoplasmic membrane and cell lysis of both Gram-positive and Gram-negative bacteria. The combination treatment also caused significant potassium and phosphate ion leakage of E. coli O157:H7, L. monocytogenes and B. thermosphacta, when compared with other treatments: 16.62±1.05, 50.35±0.81 and 45.47±1.15mg/L of potassium ion and 122.66±8.81, 97.96±3.31 and 26.47±13.97mg/L of phosphate ion after treatment for 6h, respectively. Bacteria were reduced by approximately 7log10CFU/mL within the first hour of treatment and then cells were unable to grow for the remainder of the experiment. Treatment with LAE alone resulted in changes in cellular morphology, coagulation of the cytoplasm, and low level leakage of potassium and phosphate ions in all bacteria tested. Treatment of L. monocytogenes and B. thermosphacta with nisin Z (320AU/mL of final concentration) resulted in the formation of membrane channels and leakage of potassium and phosphate ions at rather high levels; but the bacteriocin was not effective against E. coli O157:H7. LAE or nisin Z reduced growth of both L. monocytogenes and B. thermosphacta by approximately 7log10CFU/mL. Conversely, E. coli O157:H7 was not inhibited by treatments with nisin Z, but decreased by approximately 4.45log10CFU/mL after treatment with LAE. These findings provide additional information on the mode of action of these compounds on bacterial populations.

Purification and Characterization of A Novel (R)-Hydroxynitrile Lyase from Eriobotrya japonica (Loquat)

A hydroxynitrile lyase was isolated and purified to homogeneity from seeds of Eriobotrya japonica (loquat). The final yield, of 36% with 49-fold purification, was obtained by 30–80% (NH4)2SO4 fractionation and column chromatography on DEAE-Toyopearl and Concanavalin A Sepharose 4B, which suggested the presence of a carbohydrate side chain. The purified enzyme was a monomer with a molecular mass of 72 kDa as determined by gel filtration, and 62.3 kDa as determined by SDS-gel electrophoresis. The N-terminal sequence is reported. The enzyme was a flavoprotein containing FAD as a prosthetic group, and it exhibited a K m of 161 μM and a k cat⁄K m of 348 s−1 mM−1 for mandelonitrile. The optimum pH and temperature were pH 5.5 and 40 °C respectively. The enzyme showed excellent stability with regard to pH and temperature. Metal ions were not required for its activity, while activity was significantly inhibited by CuSO4, HgCl2, AgNO3, FeCl3, β-mercaptoethanol, iodoacetic acid, phenylmethylsulfonylfluoride, and diethylpyrocarbonate. The specificity constant (k cat⁄K m) of the enzyme was investigated for the first time using various aldehydes as substrates. The enzyme was active toward aromatic and aliphatic aldehydes, and showed a preference for smaller substrates over bulky one.

Incorporation of nisin Z and lauric arginate into pullulan films to inhibit foodborne pathogens associated with fresh and ready-to-eat muscle foods

A combination of food grade compounds with edible films, used to inhibit foodborne pathogens associated with fresh or further processed muscle foods, is receiving considerable attention. In this study, pullulan films containing lauric arginate (LAE) and nisin Z (produced by Lactococcus lactis subsp. lactis I8-7-3 and isolated from catfish gut), alone or in combination, were investigated for controlling foodborne pathogens on fresh and further processed muscle foods after long-term refrigerated storage. Salmonella Typhimurium and Salmonella Enteritidis on raw turkey breast slices wrapped with a film containing LAE or the combination of LAE with nisin Z were reduced throughout the experiment, 2.5 to 4.5 log10 CFU/cm(2) and 3.5 to 5.1 log10 CFU/cm(2), respectively. Film containing a combination of LAE with nisin Z reduced Staphylococcus aureus and Listeria monocytogenes Scott A inoculated onto ham surfaces by approximately 5.53 and 5.62 log10 CFU/cm(2), respectively during refrigerated storage. Escherichia coli O157:H7, O111, and O26 also were reduced by >4 log 10CFU/cm(2) on raw beef slices after treatment with the combination film and refrigerated storage. The results obtained from this study indicate the LAE- and LAE-nisin Z-containing pullulan films displayed excellent inhibition against foodborne pathogens on fresh and further processed muscle foods.

Optimization of single-cell-protein production from cassava starch using Schwanniomyces castellii

Schwanniomyces castellii B5285 grew faster and produced greater biomass and higher protein yield than either S. alluvius ATCC 26074 or S. alluvius 81Y when these amylolytic yeasts were grown with 2% (w/v) cassava starch as sole C source. With 0.5% (w/v) glutamate as N source, S. castellii reached 7.12 g cell dry mass/l, with a protein yield of 6.4 g/100 g starch. The optimal agitation speed, aeration rate and pH for growth of this yeast in a fermenter were 400 rev/min, 1.67 vol./vol.min. and 5.0, respectively. Tween 80 at 0.1% increased cell dry mass to 8.90 g/l, cell yield to 44 g/100 g starch and protein yield to 7.4 g/100 g starch.

Alternative technologies for the reduction of greenhouse gas emissions from palm oil mills in Thailand.

Alternative methodologies for the reduction of greenhouse gas (GHG) emissions from crude palm oil (CPO) production by a wet extraction mill in Thailand were developed. The production of 1 t of CPO from mills with biogas capture (four mills) and without biogas capture (two mills) in 2010 produced GHG emissions of 935 kg carbon dioxide equivalent (CO2eq), on average. Wastewater treatment plants with and without biogas capture produced GHG emissions of 64 and 47% of total GHG emission, respectively. The rest of the emissions mostly originated from the acquisition of fresh fruit bunches. The establishment of a biogas recovery system must be the first step in the reduction of GHG emissions. It could reduce GHG emissions by 373 kgCO2eq/t of CPO. The main source of GHG emission of 163 kgCO2eq/t of CPO from the mills with biogas capture was the open pond used for cooling of wastewater before it enters the biogas recovery system. The reduction of GHG emissions could be accomplished by (i) using a wastewater-dispersed unit for cooling, (ii) using a covered pond, (iii) enhancing the performance of the biogas recovery system, and (iv) changing the stabilization pond to an aerated lagoon. By using options i-iv, reductions of GHG emissions of 216, 208, 92.2, and 87.6 kgCO2eq/t of CPO, respectively, can be achieved.

Water footprints of products of oil palm plantations and palm oil mills in Thailand.

The water footprint (WF) of fresh fruit bunches (FFBs) from oil palm plantations and crude palm oil (CPO) from palm oil mills in southern and eastern Thailand were determined over 25 years. Climatic conditions, soil characteristics, and the characteristics of oil palm growth were considered. The WF of FFBs was 1063 m(3)/ton (t) on average. Green, blue, and grey waters comprised of 68, 18, and 14% of total WF, respectively. The oil palm plantations in Thailand required smaller amounts of indirect blue water. The average WF for producing a ton of CPO of seven mills was 5083 m(3). Most of the waters used in the mills originated from indirect green, blue and grey waters from the plantations. The direct blue water used in the mills had less impact on the total WF, lower than 1% of the total WF. Average percentages of green, blue, and grey waters of 69, 16, and 15% of total WF were determined for the mills, respectively. The water deprivation of the FFBs and CPO ranged from 0.73-12.9 and 3.44-58.3 m(3)H2Oeq/t, respectively. In 2013, the CPO production in Thailand including green, blue, and grey waters from plantation and blue water from mills required 11,343 million m(3) water. If the oil palm variety Suratthani 7 is used in the plantation, it would increase the yield from 15.2 to 22.8 t FFBs/ha-year and decrease the WF to 888 m(3)/t FFBs. The average value of the oil extraction rate (OER) of mills was 18.1%. With an increase in the OER of 1%, a reduction of the WF of 250 m(3)/t CPO or 5.1% of total WF could be obtained.