• Journal of Dairy Science and Biotechnology
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• v.34 no.2
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• pp.117-135
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• 2016

The present study was performed to develop a functional raw food material from hydrolyzed whey protein powder (23%-GNANA) medication containing sialic acid as a marker compound that is naturally occurring at 7% concentration in GMP (glycomacropeptide). GMP is used worldwide in foodstuffs for babies and infants and is obtained from the milk protein as safe food. While the purpose of our detailed evaluation was aimed to assess preliminary NOAEL values for and above 2,000 mg/kg/day, a clinical dose allowance for 23%-GNANA (as per characteristic of a functional health product, a highly refined test substance of 23% (v/v) sialic acid combined in GMP), at the same time we also wanted to assess the safety of GMP hydrolyzate lacking sialic acid but with identical properties as GMP. Animal safety evaluation was conducted using 23%-GNANA as the test substance, produced from hydrolyzed whey protein powder (product name: HELICOBACTROL-23; provided by Medinutrol Inc. [Korea]; composed of 23% sialic acid and GMP protein) after isolating the sialic acid using enzymes approved as food additives, with GMP as a raw material, and subsequently increasing the content of xx up to 23% through 80% (v/v) ethanol soaking and concentrating, in accordance with GLP Guideline. The animal safety evaluation mentioned above was made on the basis of toxicity in SPF Sprague-Dawley female and male rats dosed with 10 mL of the test substance diluted to 0, 1,250, 2,500, and 5,000 mg/kg directly into their stomachs for 90 d. This was determined in terms of the general symptoms and animal viability, weight and amount of feed intake, eye examination, uracrasia tests, hematological and blood biochemical disorder tests, blood coagulation test, abnormal intestine weight, abnormalities during postmortem and histopathological examinations. Statistical significance was set at P<0.05. Based on the toxicity determination, a certain minor effect associated with the test substance was observed in male rats with no major effects of the tested substance, in comparison with the control group dosed with sterilized water. Nevertheless, the NOAEL value, evaluated as per toxicity criteria, was verified as 5,000 mg/kg/day (P<0.05). Similarly, for female rats, a certain minor effect associated with the test substance was observed in 5,000 mg/kg/day dosed group, with no major effect, yet the NOAEL value (as assessed as per toxicity criteria) was determined to be 5,000 mg/kg/day (P<0.05), which was the same as for male rats. Accordingly, the NOAEL values of the test substances for all female and male rats were finally verified as 5,000 mg/kg/day (P<0.05). In conclusion, it was determined that the 23%-GNANA test substance exceeds 2,000 mg/kg/day, the clinical allowance characteristic for functional health food, and was finally evaluated to cause no safety concerns when used as a raw material in functional health food production, which was the ultimate goal of the present study.

• Preventive Nutrition and Food Science
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• v.8 no.3
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• pp.278-283
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• 2003

Textural properties of tofu manufactured with micronized full-fat soyflour (MFS) were enhanced by the addition of soy protein isolate, whey protein concentrate, chitosan oligosaccharide and mushroom powder. The MFS solution (14.2% solid content) was converted to semi-solid tofu by a two-stage heat treatment with the addition of 4% coagulant mix. The MFS tofu was evaluated by a compression test as well as sensory evaluation. To produce the semi-solid gel (MFS tofu) with reasonably high strength and toughness, the MFS solution with 14.2% solid content and 7.0% protein had to be heat treated at 121$^{\circ}C$ for 3min. The relative toughness of MFS tofu was increased by the addition of SPI, showing a 144% increase. The toughness of MFS tofu prepared with the MFS/SPI mixture was greatly increased by the addition of WPC at the level of 0.7% and the water separation from MFS tofu was greatly reduced. Furthermore, the toughness and strength of MFS/SPI tofu was enhanced by the addition of 0.1% chitosan oligosaccharide and 0.2% mushroom powder. The sensory evaluation of the tofu fortified with SPI, chitosan oligosaccharide and mushroom powder was superior to that of MFS tofu, with a higher score for overall preference.

• The Korean Journal of Food And Nutrition
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• v.3 no.2
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• pp.149-160
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• 1990

The soybean $\beta$-amylase ($\alpha$-1, 4-glucan maltohydrolase, EC 3.2.1.2) is composed of seven isozymes(I', I, II, III, IV, V and VI), and isozyme II and IV are the main components among these. The Purification of $\beta$-amylase isozymes from soybean whey were performed by ammonium sulfate fractionation, CM-Sephadex C-50 column chromatography, DEAE-Sephadex chromatography and Gel filtration. The resulted purity of $\beta$-amylase was throughly confirmed by electrophoresis, and then determined its isoelectric point and molecular weight. The results obtained were as follows, 1. Five active fractions of soybean p-amylase were derived on CM-Sephadex C-50 column chromatography. 2. Seven active bands of p-amylase isozymes were detected by isoelectric focusing gel electrophoresis, and their isoelectric points(I' to VI) were 5.07, 5.15, 5.25, 5.40, 5.55, 5.70 and 5.93, respectively. 3. Isozyme II and IV were main components of soybean $\beta$-amylase. 4. The molecular weights of both isozyme II and IV were determined to be 56,000 daltons by the result of SDS polyacrylamide gel electrophoresis. 5. Km values of main isozyme II & IV for amylopectin were determined to be 2.25 mg/ml, which suggest the same function of each isozyme.

• Journal of Dairy Science and Biotechnology
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• v.34 no.2
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• pp.99-116
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• 2016

We herein performed animal safety assessment in accordance with Good Laboratory Practice (GLP) regulations with the aim of developing sialic acid from glycomacropeptide (hereafter referred to as "GMP") as an index ingredient and functional component in functional foods. GMP is a type of whey protein derived from milk and a safe food, with multiple functions, such as antiviral activity. A test substance was produced containing 7% (w/w) sialic acid and mostly-hydrolyzed whey protein (hereafter referred to as "7%-GNANA") by enzymatic treatment of substrate GMP. The maximum intake test dose level was selected based on 5,000 mg/kg/day dose set for male NOEL (no-observed-effect-level) and female NOAEL (no-observed-adverse-effect-level) determined by a dose-range finding (DRF) test (GLP Center of Catholic University of Daegu, Report No. 15-NREO-001) that was previously conducted with the same test substance. To evaluate the toxicity of a repeated oral dose of the test substance in connection with the previous DRF study, 1,250, 2,500, and 5,000 mg/kg of the substance were administered by a probe into the stomachs of 6-week-old SPF Sprague-Dawley male and female rats for 90 d. Each test group consisted of 10 male and 10 female rats. To determine the toxicity index, all parameters, such as observation of common signs; measurements of body weight and food consumption; ophthalmic examination; urinalysis, electrolyte, hematological, and serum biochemical examination; measurement of organ weights during autopsy; and visual and histopathological examinations were conducted according to GLP standards. After evaluating the results based on the test toxicity assessment criteria, it was determined that NOAEL of the test substance, 7%-GNANA, was 5,000 mg/kg/day, for both male and female rats. No animal death was noted in any of the test groups, including the control group, during the study period, and there was no significant difference associated with test substance, as compared with the control group, with respect to general symptoms, body weight changes, food consumption, ophthalmic examination, urinalysis, hematological and serum biochemical examination, and electrolyte and blood coagulation tests during the administration period (P<0.05). As assessed by the effects of the test substance on organ weights, food consumption, autopsy, and histopathological safety, change in kidney weight as an indicator of male NOAEL revealed up to 20% kidney weight increase in the high-dose group (5,000 mg/kg/day) compared with the change in the control group. However, it was concluded that this effect of the test substance was minor. In the case of female rats, reduction of food consumption, increase of kidney weight, and decrease of thymus weight were observed in the high-dose group. The kidney weight increased by 10.2% (left) and 8.9% (right) in the high-dose group, with a slight dose-dependency compared with that of the control group. It was observed that the thymus weight decreased by 25.3% in the high-dose group, but it was a minor test substance-associated effect. During the autopsy, botryoid tumor was detected on the ribs of one subject in the high-dose group, but we concluded that the tumor has been caused by a naturally occurring (non-test) substance. Histopathological examination revealed lesions on the kidney, liver, spleen, and other organs in the low-dose test group. Since these lesions were considered a separate phenomenon, or naturally occurring and associated with aging, it was checked whether any target organ showed clear symptoms caused by the test substance. In conclusion, different concentrations of the test substance were fed to rats and, consequently, it was verified that only a minor effect was associated with the test substance in the high-dose (5,000 mg/kg/day) group of both male and female rats, without any other significant effects associated with the test substance. Therefore, it was concluded that NOAEL of 7%-GNANA (product name: Helicobactrol) with male and female rats as test animals was 5,000 mg/kg/day, and it thus was determined that the substance is safe for the ultimate use as an ingredient of health functional foods.

• Journal of Applied Biological Chemistry
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• v.59 no.4
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• pp.345-349
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• 2016

This study was performed to evaluate the quality characteristics of curd yogurt with different contents [0.5~2.0% (w/w)] of Korean rice wine lees powder (KRWLP). Yogurt was fermented with commercially available mixed lactic acid bacteria (Lactobacillus acidophilus, Bifidobacterium longum, Streptococcus thermophiles) at $40^{\circ}C$ for 15 h. Acid production (pH and titratable acidity) of yogurts increased with increasing KRWLP content. After 12 hours fermentation, titratable acidity of KRWLP yogurt was 1.19~1.29 % and was higher than that (1.07 %) of yogurt made without KRWLP. And also, the number of viable lactic acid bacterial cell increased and the culture time to obtain maximum number of lactic acid bacteria cell decreased with the addition of KRWLP. The curd stability in yogurt was significantly enhanced by repression of whey separation in KRWLP yogurt. In sensory evaluation, there was a similar preference for KRWLP yogurts and the control. These results suggest that KRWLP can be used as foodstuff to improve the quality characteristics of yogurt.

• Food Science of Animal Resources
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• v.39 no.1
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• pp.23-34
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• 2019

Fat reduction in the formulation of cheese emulsion causes problems in its flowability and functional characteristics during spray-dried cheese powder production. In order to eliminate these problems, the potential of using microparticulated whey protein (MWP) in cheese emulsions was examined in this study. Reduced-fat white-brined cheese emulsions (RF) with different dry-matters (DM) (15%, 20%, and 25% excluding emulsifying salt) were produced using various MWP concentrations (0%-20% based on cheese DM of emulsion). Their key characteristics were compared to full-fat cheese emulsion (FF). MWP addition had no influence on prevention of the phase separation observed in the instable group (RF 15). The most notable effect of using MWP was a reduction in apparent viscosity of RF which significantly increased by fat reduction. Moreover, increasing the amount of MWP led to a decrease in the values of consistency index and an increase in the values of flow behavior index. On the other hand, using high amounts of MWP made the emulsion more liquid-like compared to full-fat counterpart. MWP utilization also resulted in similar lightness and yellowness parameters in RF as their full-fat counterparts. MWP in RF increased glossiness and flowability scores, while decreased mouth coating scores in sensory analyses. Fat reduction caused a more compact network, while a porous structure similar to FF was observed with MWP addition to RF. In conclusion, MWP showed a good potential for formulation of reduced-fat cheese emulsions with rheological and sensorial characteristics suitable to be used as the feeding liquid in the spray drying process.

• Journal of Dairy Science and Biotechnology
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• v.34 no.2
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• pp.91-98
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• 2016

The goal of this study was to develop hydrolyzed whey protein powder (23%-GNANA) manufactured with high content of sialic acid, a marker compound that is usually present at 7% concentration in GMP obtained from the milk protein. It is a safe food, used worldwide in infant and baby foods, etc. The test substance was prepared using (7% sialic acid containing) GMP as a raw material. Alcalase, an enzyme approved as a food additive, was used after separating sialic acid, with 100% efficiency, and 23%-GNANA (composed of 23% sialic acid and protein; product name: HELICOBACTROL-23), provided by MEDINUTROL Inc. (Korea), manufactured to have high (23%) content through ethanol soaking and enrichment. Bacterial reverse mutation (Ames) test was conducted in accordance with the GLP Guideline using the test substance specified above. To detect its mutagenicity potential in microorganisms, histidine auxotrophic strains of Salmonella typhimurium, TA98, TA100, TA1535, and TA1537, and tryptophan auxotrophic Escherichia coli strain, WP2uvrA, were used. The bacterial reverse mutation (Ames) test was performed using five concentrations of the test substances (0, 61.7, 185, 556, 1,670, $5,000{\mu}g/plate$). The evaluation did not reveal repetitive increase of colony generating values and positive criteria for reverse mutagenicity for any tested concentration in the five strains regardless of the presence of metabolic activation system, and no dose-dependency. In conclusion, the safety of 23%-GNANA test substance was verified by the bacterial reverse mutation test conducted before registration of 23%-GNANA as a food additive.

• Journal of Dairy Science and Biotechnology
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• v.34 no.2
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• pp.83-89
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• 2016

This study was designed to determine the mutagenic potential of hydrolyzed glycomacropeptide (GMP) powder (hereafter referred to as 23%-GNANA; product name: HELICOBACTROL-23) in a micronucleus test using bone marrow in ICR mice. Three experimental groups were used: a 3-step concentration group, with a maximum concentration of 2,000 mg/kg, and other sequentially two-fold lower concentrations, a negative control group, and a positive control group. The test material was administered for 2 d to observe the frequency of micronucleus formation up to 48 h after the test material was absorbed by the body. When the polychromatic erythrocyte (PCE) content of erythrocytes was compared, no significant differences were noted between the negative control group and the test group (p<0.05). Similarly, when the average numbers of micronucleated PCE (MNPCE) in 2,000 PCE per animal were compared, no significant difference was observed between the negative control group and the test group (p<0.05). No dose-response relationship with regard to the concentration of the test material administered was noted. These results allow us to conclude that hydrolyzed whey protein powder does not cause formation of micronuclei in mouse bone marrow cells under the applied conditions. In this study, the average frequency of micronucleus formation in PCE was significantly higher in the positive control group compared with the negative control group; thus, the test conditions were appropriate for detecting the frequency of micronucleus formation induced by the test material. In conclusion, the safety of 23%-GNANA test substance was verified in an in vivo micronucleus test in mice, conducted before the registration of HELICOBACTROL-23 as a food additive.

• Proceedings of the Membrane Society of Korea Conference
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• 1998.06a
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• pp.135-153
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• 1998

Nanofiltration (NF) is a recently introduced term in membrane separation. In 1988, Eriksson was one of the first authors using the word 'nanofiltration' explicitly. Some years before, FilmTech started to use this term for their NF50 membrane which was supposed to be a very loose reverse osmosis membrane or a very tight ultrafiltration membrane. Since then, this term has been introduced to indicate a specific boundary of membrane technology in between ultrafiltration and reverse osmosis. The application fields of the NF membranes are very broad as follows: Demeneralizing water, Cleaning up contaminated groundwater, Ultrapure water production, Treatment of effleunts containing heavy metals, Offshore oil platforms, Yeast production, Pulp and paper mills, Textile production, Electroless copper plating, Cheese whey production, Cyclodextrin production, Lactose production. The earliest NF membrane was made by Cadotte et al, using piperazine and trimesoyl chloride as monomers for the formation of polyamide active layer of the composite type membrane. They coated very thin interfacially potymerized polyamide on the surface of the microporous polysulfone supports. The NF membrane exhibited low rejections for monovalent anions (chloride) and high rejections for bivalent anions (sulphate). This membrane was called NS300. Some of the earliest NF membranes, like the NF40 membrane of FilmTech, the NTR7250 of Nitto-Denko and the UTC20 and UTC60 of Toray, are formed by a comparable synthesis route as the NS300 membrane. Commercially available NF membranes nowadays are as follows: ASP35 (Advanced Membrane Technology), MPF21; MPF32 (Kiryat Weizmann), UTC20; UTC60; UTC70; UTC90 (Toray), CTA-LP; TFCS (Fluid Systems), NF45; NF70 (FilmTec), BQ01; MX07; HG01; HG19; SX01; SX10 (Osmonics), 8040-LSY-PVDI (Hydranautics), NF CA30; NF PES 10 (Hoechst), WFN0505 (Stork Friesland). The typical ones among the commercially available NF membranes are polyamide composite membrane consisting of interfacially polymerized polyamide active layer and microporous support. While showing high water fluxes and high rejections of multivalent ions and small organic molecules, these membranes have relatively low chemical stability. These membranes have low chlorine tolerance and are unstable in acid or base solution. This chemical instability is appearing to be a big obstacle for their applications. To improve the chemical stability, we have tried, in this study, to prepare chemically stable NF membranes from PVA. The ionomers and interfacially polymerized polyamide were used for the modification of'the PVA membranes. For the detail study of the active layer, homogeneous NF membranes made only from active layer materials were prepared and for the high performance, composite type NF membranes were prepared by coating the active layer materials on microporous polysulfone supports.

• Korean Journal of Food Science and Technology
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• v.46 no.2
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• pp.165-172
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• 2014

Mixed fermentation of cow milk was performed by sequential co-cultures with Bacillus subtilis and Lactococcus lactis. After a first fermentation step with B. subtilis for 6 h, the number of viable cells increased to $2.5{\times}10^8$ CFU/mL. The second fermentation step with L. lactis resulted in increased viable cells $1.09{\times}10^{10}$ CFU/mL for 3 days and increased acidity. However, the number of viable B. subtilis cells was decreased greatly to $5{\times}10^1$ CFU/mL following fermentation with L. lactis. Milk proteins were markedly hydrolyzed by the first fermentation after 2 h, and the second fermentation induced curd formation in milk. However, after 4 h, the first fermentation resulted in higher whey separation and 80 mg% tyrosine content. Gamma-aminobutyric acid (GABA) production was dependent upon the degree of protein hydrolysis by first fermentation. Second fermentation resulted in 0.14% GABA. The milk fermented by B. subtilis indicated the rough surface of yogurt depended upon the degree of protein hydrolysis. In conclusion, set-type yogurt was efficiently produced by co-culturing of milk, and fortifying with peptides, GABA, and probiotics.