• Korean Journal of Food Science and Technology
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• v.2 no.2
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• pp.38-42
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• 1970

The crude enzyme, tamylase, was produced by cultivating the Bacillus subtilis on wheat bran. It is composed of amylase and protease, and can be used as an additive material to the synthetic detergent, Suny which is manufactured by Ae-kyung Oil and Fat Co. Amylase activity of the enzyme as an additive material to the synthetic detergent; 1. is decreased by increasing the amount of detergent. But inhibitory rate under the practical used concentration of detergent is less than ten percents. 2. have optimal temperature at $ 40^{\circ}C$. 3. have optimal pH of substrate on pH $5{\sim}6.5$. 4. is inhibited by $Fe^{+++}$. When enzyme and detergent are mixed both as powder, the enzyme is good for storage. Proteolytic activity is good at the practical used concentration of the detergent, but it is inhibited by strong concentration.

• Journal of Microbiology and Biotechnology
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• v.33 no.2
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• pp.195-202
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• 2023

Protease is a widely used enzyme particularly in the detergent industry. In this research, we aimed to isolate alkaline protease-producing bacteria for characterization as a laundry detergent additive. The screening of alkaline protease production was investigated on basal medium agar plus 1% skim milk at pH 11, with incubation at 30℃. The highest alkaline protease-producing bacterium was 6BS15-4 strain, identified as Bacillus gibsonii by 16S rRNA gene sequencing. While the optimum pH was 12.0, the strain was stable at pH range 7.0-12.0 when incubated at 45℃ for 60 min. The alkaline protease produced by B. gibsonii 6BS15-4 using dairy effluent was characterized. The optimum temperature was 60℃ and the enzyme was stable at 55℃ when incubated at pH 11.0 for 60 min. Metal ions K⁺, Mg²⁺, Cu²⁺, Na⁺, and Zn²⁺ exhibited a slightly stimulatory effect on enzyme activity. The enzyme retained over 80% of its activity in the presence of Ca²⁺, Ba²⁺, and Mn²⁺. Thiol reagent and ethylenediaminetetraacetic acid did not inhibit the enzyme activity, whereas phenylmethylsulfonyl fluoride significantly inhibited the protease activity. The alkaline protease from B. gibsonii 6BS15-4 demonstrated efficiency in blood stain removal and could therefore be used as a detergent additive, with potential for various other industrial applications.

• Asian-Australasian Journal of Animal Sciences
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• v.17 no.7
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• pp.942-946
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• 2004

This study aimed to examine the fermentation quality of a tropical Elephant grass (Pennisetum purpuereum) and temperate Italian ryegrass (Lolium multiflorum) forages ensiled additive of fermented juice of epiphytic lactic acid bacteria (LAB) and to determine what factor affects the fermentation characteristics of the crops. In both species cell walls neutral detergent fiber (NDF) and acid detergent fiber (ADF) contents in silages were consistently decreased (p<0.05) with the addition of FJLB at ensiling more then Control treatment. The FJLB additive increased number of LAB (cfu) and lactate concentration in the silages in both species. The Control treatment without additive underwent a clostridial type of fermentation with traces of propionic, iso-butyric, n-butyric acids contents with higher (p<0.01) levels of volatile basic nitrogen (VBN %TN) and had appreciable decreased of nutrient in silages. FJLB treatment improved silage nutritive value with little contents of VBN %TN, ethanol and very small amount of dry matter (DM) and hemicellulose losses (p<0.05) between 2 to 5% and 7 to 3% respectively, in Elephant grass and Italian ryegrass species. The results in this study indicates that while among the factors affecting silage fermentation butyric type of fermentation was more pronounced in tropical elephant grass compared to the temperate Italian ryegrass, FJLB additive revealed a better silage fermentation products in both species.

• Microbiology and Biotechnology Letters
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• v.47 no.2
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• pp.259-268
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• 2019

In this study, the extracellular metalloprotease from Serratia marcescens PPB-26 was purified to homogeneity via ethanol fractionation and DEAE-cellulose column chromatography. Thus, a 3.8-fold purification was achieved with a 20% yield and specific activity of 76.2 U/mg. The purified protease was a 50-kDa monomer whose optimum pH and temperature for activity were 7.5 and $30^{\circ}C$ respectively; however, it was found to remain active in the 5-9 pH range and up to $40^{\circ}C$ for 6 h. The protease had a half-life of 15 days at $4^{\circ}C$, an optimum reaction time of 10 min, and an optimum substrate (casein) concentration of 0.25%. Furthermore, the Michaelis constant ($K_m$) and reaction velocity ($V_{max}$) of the protease were calculated to be 0.28% and $111.11{\mu}moles/(min{\cdot}mg)^{-1}$, respectively. The protease was stable when subjected to metal ions (2 mM), showing increased activity with most (especially $CoCl_2$ and $MgSO_4$ (30.54% increase)). It was also stable when exposed to oxidizing agents, bleaching agents, and detergents (5% v/v for 60 min). It retained 93% of its activity in non-ionic detergents (Tween-20, Tween-80, and Triton X-100). Moreover, wash performance analysis in commercial detergents (Ariel and Tide) showed that not only was the protease capable of protein stain removal, but also reduced cleaning time by 80% when added to detergents. Thus, the Serratia marcescens PPB-26 metalloprotease appears to be a promising new candidate as a laundry additive in the detergent industry.

• Microbiology and Biotechnology Letters
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• v.48 no.3
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• pp.358-365
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• 2020

An organic solvent- and bleach-stable protease-producing strain was isolated from a polluted river water sample and identified as Aeromonas veronii OB3 on the basis of biochemical properties (API 20E) and 16S rRNA sequence analysis. The strain was found to hyper-produce alkaline protease when cultivated on fish waste powder-based medium (HVSP, 4080 U/ml). The biochemical properties and compatibility of OB3 with several detergents and additives were studied. Maximum activity was observed at pH 9.0 and 60℃. The crude protease displayed outstanding stability to the investigated surfactants and oxidants, such as Tween 80, Triton X-100, and H₂O₂, and almost 36% residual activity when incubated with 1% SDS. Remarkably, the enzyme demonstrated considerable compatibility with commercial detergents, retaining more than 100% of its activity with Ariel and Tide (1 h, 40℃). Moreover, washing performance of Tide significantly improved by the supplementation of small amounts of OB3 crude protease. These properties suggest the potential use of this alkaline protease as a bio-additive in the detergent industry and other biotechnological processes such as peptide synthesis.

• Animal Bioscience
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• v.34 no.1
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• pp.48-55
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• 2021

Objective: This study was conducted to investigate the effects of wilting and microbial inoculant treatment on the fermentation pattern and quality of Italian ryegrass silage. Methods: Italian ryegrass was harvested at heading stage and ensiled into vinyl bags (20 cm×30 cm) for 60d. Italian ryegrass was ensiled with 4 treatments (NWNA, no-wilting noadditive; NWA, no-wilting with additive; WNA, wilting no-additive; WA, wilting with additive) in 3 replications, wilting time was 5 hours and additives were treated with 106 cfu/g of Lactobacillus plantarum. The silages samples were collected at 1, 2, 3, 5, 10, 20, 30, 45, and 60 days after ensiling and analyzed for the ensiling quality and characteristics of fermentation patterns. Results: Wilting treatment resulted in lower crude protein and in vitro dry matter digestibility and there were no significant differences in acid detergent fiber (ADF), total digestible nutrient (TDN), water-soluble carbohydrate (WSC), ammonia content, and pH (p>0.05). However, wilting treatment resulted in higher ADF and neutral detergent fiber content of Italian ryegrass silage (p<0.05), and the WNA treatment showed the lowest TDN and in vitro dry matter digestibility. The pH of the silage was higher in the wilting group (WNA and WA) and lower in the additive treatment group. Meanwhile, the decrease in pH occurred sharply between the 3-5th day of storage. The ammonia nitrogen content was significantly lower in the additive treatment (p<0.05), and wilting had no effect. As fermentation progressed, the lactic and acetic acid contents were increased and showed the highest content at 30 days of storage. Conclusion: The wilting treatment did not significantly improve the silage fermentation, but the inoculant treatment improved the fermentation patterns and quality of the silage. So, inoculation before ensiling is recommended when preparing high quality of Italian ryegrass silage, and when wilting, it is recommended to combine inoculation for making high quality silage.

• KSBB Journal
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• v.27 no.6
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• pp.352-360
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• 2012

Bacillus clausii I-52 which produced SDS- and $H_2O_2$-tolerant extracellular alkaline protease (BCAP) was isolated from heavily polluted tidal mud flat of West Sea in Incheon, Korea and stable strain (transformant C5) of B. clausii I-52 harboring another copy of BCAP gene in the chromosome was developed using the chromosome integration vector, pHPS9-fuBCAP. When investigated the production of BCAP using B. clausii transformant C5 through pilot-scale submerged fermentation (500 L) at $37^{\circ}C$ for 30 h with an aeration rate of 1 vvm and agitation rate of 250 rpm, protease yield of approximately 105,700 U/mL was achieved using an optimized medium (soybean meal 2%, wheat flour 1%, sodium citrate 0.5%, $K_2HPO_4$ 0.4%, $Na_2HPO_4$ 0.1%, NaCl 0.4%, $MgSO_4{\cdot}7H_2O$ 0.01%, $FeSO_4{\cdot}7H_2O$ 0.05%, liquid maltose 2.5%, $Na_2CO_3$ 0.6%). The enzyme stability of BCAP was increased by addition of polyols (10%, v/v) and also, the stabilities of BCAP towards not only the thermal-induced inactivation at $50^{\circ}C$ but also the SDS and $H_2O_2$-induced inactivation at $50^{\circ}C$ were enhanced. Among the polyols examined, the best result was obtained with propylene glycol (10%, v/v). The BCAP supplemented with propylene glycol exhibited extreme stability against not only the detergent components such as ${\alpha}$-orephin sulfonate (AOS) and zeolite but also the commercial detergent preparations. The granulized enzyme of BCAP was prepared with approximately 1,310,000 U/g of granule. Wash performance analysis using EMPA test fabrics revealed that BCAP granule exhibited high efficiency for removal of protein stains in the presence of anionic surfactants as well as bleaching agents. When compared to Savinase 6T$^{(R)}$ and Everlase 6T$^{(R)}$ manufactured by Novozymes, BCAP under this study probably showed similar or higher efficiency for the removal of protein stains. These results suggest that the alkaline protease produced from B. clausii transformant C5 showing high stability against detergents and high wash performance has significant potential and a promising candidate for use as a detergent additive.

• Asian-Australasian Journal of Animal Sciences
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• v.33 no.2
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• pp.245-253
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• 2020

Objective: To develop the fermentation quality and chemical composition of alfalfa (Medicago sativa Lam.) silage, plants were inoculated with different lactic acid bacteria (LAB) strains at field 24 hours before harvest. Methods: The treatment groups were as follow: silage without additive as a control and inoculated with each strains of Lactobacillus brevis (LS-55-2-2), Leuconostoc citerum (L. citerum; L-70-6-1), Lactobacillus bifermentans (L. bifermentans; LS-65-2-1), Lactobacillus plantarum (L. plantarum; LS-3-3) and L. plantarum (LS-72-2). All the silages were stored at 25℃. Parameters such as pH, microorganism and volatile fatty acid contents, crude protein, neutral detergent fiber, acid detergent fiber, net gas, metabolizable energy, organic matter digestibility, dry matter intake and relative feed value were measured to determine fermentation quality, chemical compositions and relative feed value of alfalfa silages. Results: Significant differences were found among the control and treated groups in terms of pH and microorganism contents at all opening times and crude protein, net gas, metabolizable energy and organic matter digestibility of final silage. The pH values ranged from 4.70 to 5.52 for all treatments and control silage had the highest value of overall treatments at T₇₅d silages. Volatile fatty acid of silages was not influenced significantly by inoculations. However, lactic acid content of L. bifermentans (LS-65-2-1) was higher than the other treatments. The highest metabolizable energy and organic matter digestibility were recorded from L. citerum (L-70-6-1) inoculation. In addition, no significant differences were found among treatments in terms of neutral detergent fiber, acid detergent fiber, dry matter intake and relative feed value. Conclusion: Among the treated LAB isolates, L. bifermentans came into prominence especially in terms of organic acid composition and quality characters of silages.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.5
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• pp.672-676
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• 2018

Objective: An experiment was conducted to study the effect of a blend of essential oils (BEO) on enteric methane emission and growth performance of buffaloes (Bubalus bubalis). Methods: Twenty one growing male buffaloes (average body weight of $279{\pm}9.3kg$) were divided in to three groups. The animals of all the three groups were fed on a ration consisting of wheat straw and concentrate mixture targeting 500 g daily live weight gain. The three dietary groups were; Group 1, control without additive; Group 2 and 3, supplemented with BEO at 0.15 and 0.30 mL/kg of dry matter intake (DMI), respectively. Results: During six months feeding trial, the intake and digestibility of dry matter and nutrients (organic matter, crude protein, ether extract, neutral detergent fibre, and acid detergent fibre) were similar in all the groups. The average body weight gain was tended to improve (p = 0.084) in Group 2 and Group 3 as compared to control animals. Feeding of BEO did not affect feed conversion efficiency of the animals. The calves of all the three groups were in positive nitrogen balance with no difference in nitrogen metabolism. During respiration chamber studies the methane production (L/kg DMI and L/kg digestible dry matter intake was significantly (p<0.001) lower in Group 2 and Group 3 as compared to control animals. Conclusion: The results indicated that the BEO tested in the present study have shown potential to reduce enteric methane production without compromising the nutrient utilization and animal performance and could be further explored for its use as feed additive to mitigate enteric methane production in livestock.

• Asian-Australasian Journal of Animal Sciences
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• v.25 no.9
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• pp.1248-1254
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• 2012

The effect of silage additives on ensiling characteristics and nutritive value of Napier grass (Pennisetum purpureum) silages was studied. Napier grass silages were made with no additive, fermented juice of epiphytic lactic acid bacteria (FJLB), molasses or cassava meal. The ensiling characteristics were determined by ensiling Napier grass silages in airtight plastic pouches for 2, 4, 7, 14, 21 and 45 d. The effect of Napier grass silages treated with these additives on voluntary feed intake, digestibility, rumen fermentation and microbial rumen fermentation was determined in 4 fistulated cows using $4{\times}4$ Latin square design. The pH value of the treated silages rapidly decreased, and reached to the lowest value within 7 d of the start of fermentation, as compared to the control. Lactic acid content of silages treated with FJLB was stable at 14 d of fermentation and constant until 45 d of ensiling. At 45 d of ensiling, neutral detergent fiber (NDF) and acid detergent fiber (ADF) of silage treated with cassava meal were significantly lower (p<0.05) than the others. In the feeding trial, the intake of silage increased (p<0.05) in the cow fed with the treated silage. Among the treatments, dry matter intake was the lowest in the silage treated with cassava meal. The organic matter, crude protein and NDF digestibility of the silage treated with molasses was higher than the silage without additive and the silage treated with FJLB. The rumen parameters: ruminal pH, ammonia-nitrogen ($NH_3$-N), volatile fatty acid (VFA), blood urea nitrogen (BUN) and bacterial populations were not significantly different among the treatments. In conclusion, these studies confirmed that the applying of molasses improved fermentative quality, feed intake and digestibility of Napier grass.