• Archives of Pharmacal Research
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• 제26권9호
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• pp.731-734
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• 2003

Five known kaurane type diterpenoids, 16$\alpha$H, 17-isovaleryloxy-ent-kauran-19-oic acid (1), 16$\alpha$-hydroxy-17-isovaleryloxy-ent-kauran-19-oic acid (2), paniculoside-IV (3), 16$\alpha$-hydroxy-ent-kauran-19-oic acid (4), and ent-kaur-16-en-19-oic acid (5) were isolated from the root of Acanthopanax koreanum by repeated column chromatography and reversed phase preparative HPLC. The structures of these compounds were established from physicochemical and spectral data. Among the isolated compounds 16$\alpha$H, 17-isovaleryloxy-ent-kauran-19-oic acid (1) showed potent inhibitory activity ($IC_50$ value, 16.2 $\mu$ M) on TNF-$\alpha$ secretion from HMC-1, a trypsin-stimulated human leukemic mast cell line.

• Fisheries and Aquatic Sciences
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• 제21권5호
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• pp.13.1-13.8
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• 2018

Amyloid plaque, also called senile plaque, the product of aggregation of ${\beta}$-amyloid peptides ($A{\beta}$), is observed in brains of the patients with Alzheimer's disease (AD) and is one of the key factors in etiology of the disease. In this study, hydrolysates obtained from the sea hare (Aplysia kurodai) were investigated for ${\beta}$-secretase inhibitory peptide. The sea hare's muscle protein was hydrolyzed using six enzymes in a batch reactor. Trypsin hydrolysate had highest ${\beta}$-secretase inhibitory activity compared to the other hydrolysates. ${\beta}$-secretase inhibitory peptide was separated using Sephadex G-25 column chromatography and high-performance liquid chromatography on a C18 column. ${\beta}$-secretase inhibitory peptide was identified as eight amino acid residues of Val-Ala-Ala-Leu-Met-Leu-Phe-Asn by N-terminal amino acid sequence analysis. $IC_{50}$ value of purified ${\beta}$-secretase inhibitory peptide was $74.25{\mu}M$, and Lineweaver-Burk plots suggested that the peptide purified from sea hare muscle protein acts as a competitive inhibitor against ${\beta}$-secretase. Results of this study suggest that peptides derived from sea hare muscle may be beneficial as anti-dementia compounds in functional foods or as pharmaceuticals.

• 한국식품과학회지
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• 제26권4호
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• pp.464-470
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• 1994

환삼덩굴의 생리활성물질을 검색하기 위하여 물과 메탄올로 추출하여 항균성과 trypsin 저해활성 및 항산화 효과를 조사하였으며, 메탄올 추출물은 다시 계통 분획하여 분획별 활성을 조사하였다. 세균 6종과 효모 2종을 대상으로 항균성을 조사한 결과, 물 추출물 보다는 메탄올 추출물의 항균성이 우수하였으며, Escherichia coli를 제외한 모든 시험균주에 항균효과를 나타내었다. 분획별 항균효과를 비교한 결과, 부탄올 분획이 모든 시험균주에 항균효과를 나타내었으며, 균종별로는 Staphylococcus aureus와 Saccharomyces cerevisiae에 대한 항균효과가 우수하였다. 부탄올 분획의 미생물에 대한 최소저해농도(MIC)는 $0.1{\sim}0.4%$ 범위였다. 환삼덩굴 추출물의 trypsin 저해활성을 조사한 결과, 물 추출물은 저해활성을 나타내지 않았으나, 메탄올 추출물은 저해활성을 나타내었다. 분획별 저해활성은 클로로포름 분획에서 가장 높았으며 클로로포름 분획의 50% 저해농도($IC_{50}$)는 1mg/ml였으며, EI(enzyme-inhibitor) 복합체 형성시간은 20분에 90% 이상이었다. 환삼덩굴의 메탄올 추출물은 돈지와 대두유를 기질로한 항산화성 시험에서 과산화물의 생성을 억제하였다. 메탄올 추출물의 분획별 활성은 클로로포름 분획에서 가장 높게 나타났으며, 클로로포름 분획의 농도를 0.2% 첨가하였을 때, 유지의 유도기간은 돈지는 15일, 대두유는 9일로서 산화안정성이 연장되었다.

• Natural Product Sciences
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• 제26권3호
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• pp.207-213
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• 2020

Characterization and in vitro inhibition studies of protease inhibitor from the mushroom Pleurotus floridanus (PfTI) towards the pest Papilio demoleus is studied. The addition of 1 mM Mn²⁺, Na²⁺, Ba²⁺ and Ni ²⁺ enhanced the PfTI activity. The ICP-atomic emission spectrum showed the presence of Ca²⁺, Mg²⁺ and Zn²⁺ in the PfTI. Surfactants SDS and CTAB at a concentration of 1% reduced the PfTI activity whereas, the nonionic detergents Triton X and Tween 80 increased the activity. The inhibitory activity gradually decreased with increase in concentration of DMSO and H₂O₂. The activity was increased by dithiothreitol up to a concentration of 80 μM and inactivated at 140 μM. The activity of PMSF modified PfTI was drastically reduced to 0.234 U/mL at 4 mM concentration and similar results were obtained for modification of cysteine by N-Ethylmaleimide at slightly higher concentrations. The complex of trypsin and PfTI showed complete loss in fluorescence intensity at 343 nm compared with control. In vitro inhibition studies of PfTI with midgut proteases isolated from citrus pest P. demoleus with protease activity of 1.236 U was decreased to 0.613 U by 50 μL (0.1 mg/mL) of the inhibitor. Inhibitor was stable up to 0.04 M concentration of HCl.

• 한국수산과학회지
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• 제39권3호
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• pp.269-277
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• 2006

This study prepared functional oyster hydrolysates using two-step enzymatic hydrolysis and investigated their functional properties. To prepare two-step enzymatic hydrolysates (TSEH), oysters were hydrolyzed using 1% Protamex (PR) at $40^{\circ}C$ and pH 6.0 for 1 hr before sequential treatment with one of the following enzymes for 1 hr: Alcalase (AL), Flavourzyme (FL), Neutrase (NE), pepsin (PE), and trypsin (TR). The PRAL, PRNE and PRTR hydrolysates had significantly greater angiotensin I converting enzyme (ACE) inhibitory activity than did PR and the other TSEHs. Only the antioxidant activity of the PRNE hydrolysate was significantly different (p<0.05), while none of the TSEHs had antimicrobial activity. The oyster hydrolysate prepared by sequential treatment with Protamex and Neutrase (PRNE) had the best ACE inhibitory activity and antioxidant activity, with $IC_{50}$ values of 0.40 and 0.94 mg/mL, respectively. The PRNE hydrolysate was processed through an ultrafiltration (UF) series with molecular weight cut-off (MWCO) membranes of 3, 5, 10, and 30 kDa, and the ACE inhibitory, antioxidant, and antimicrobial activities of the permeates were determined. The permeate through the 3-kDa MWCO membrane had greater ACE inhibitory activity and antioxidant activity than did the other PRNE permeates, with $IC_{50}$ values of 0.11 and 0.40 mg/mL, respectively.

• Food Science and Biotechnology
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• 제17권3호
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• pp.489-494
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• 2008

In order to develop a potent antidementia $\beta$-secretase inhibitor from phytochemicals, $\beta$-secretase inhibitory activities of extracts from many medicinal plants and herbs were determined. Water extracts from Rubus coreanus showed the highest $\beta$-secretase inhibitory activity of 84.5%. After purification of the $\beta$-secretase inhibitor from R. coreanus using systematic solvent extraction, ultrafiltration, Sephadex G-10 column chromatography, and reverse-phase high performance liquid chromatography (HPLC), a purified $\beta$-secretase inhibitor with $IC_{50}$ inhibitory activity of $6.3{\times}10^3\;ng/mL$ ($1.56{\times}10^{-6}\;M)$ was obtained with a 0.08% solid yield. The molecular mass of the purified $\beta$-secretase inhibitor was estimated to be 576 Da by liquid chromatography-mass spectrometry (LC-MS) and $\beta$-secretase inhibitor also is a new tetrapeptide with the sequence Gly-Trp-Trp-Glu. The purified $\beta$-secretase inhibitory peptide inhibited $\beta$-secretase non-competitively and also show less inhibition on trypsin, however no inhibition on other proteases such as $\alpha$-secretase, chymotrypsin, and elastase.

• Journal of Microbiology and Biotechnology
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• 제28권12호
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• pp.2079-2094
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• 2018

Sunflower trypsin inhibitor (SFTI) is a 14-amino-acid bicyclic peptide that contains a single internal disulfide bond. We initially constructed chimeras of SFTI with N-terminal secretion signals from the Escherichia coli OmpA and Pseudomonas aeruginosa ToxA, but only detected small amounts of protease inhibition resulting from these constructs. A substantially higher degree of protease inhibition was detected from a C-terminal SFTI fusion with E. coli YebF, which radiated more than a centimeter from an individual colony of E. coli using a culture-based inhibitor assay. Inhibitory activity was further improved in YebF-SFTI fusions by the addition of a trypsin cleavage signal immediately upstream of SFTI, and resulted in production of a 14-amino-acid, disulfide-bonded SFTI free in the culture supernatant. To assess the potential of the secreted SFTI to protect the ability of a cytotoxic protein to kill tumor cells, we utilized a tumor-selective form of the Pseudomonas ToxA (OTG-PE38K) alone and expressed as a polycistronic construct with YebF-SFTI in the tumor-targeted Salmonella VNP20009. When we assessed the ability of toxin-containing culture supernatants to kill MDA-MB-468 breast cancer cells, the untreated OTG-PE38K was able to eliminate all detectable tumor cells, while pretreatment with trypsin resulted in the complete loss of anticancer cytotoxicity. However, when OTG-PE38K was co-expressed with YebF-SFTI, cytotoxicity was completely retained in the presence of trypsin. These data demonstrate SFTI chimeras are secreted in a functional form and that co-expression of protease inhibitors with therapeutic proteins by tumor-targeted bacteria has the potential to enhance the activity of therapeutic proteins by suppressing their degradation within a proteolytic environment.

• Journal of Microbiology and Biotechnology
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• 제10권2호
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• pp.181-186
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• 2000

MAPI (microbial alkaline protease inhibitor) was isolated from cultrue broth of Streptomyces chromofuscus SMF28. The Ki values of MAPI for the representative serine proteases such as chymotrypsin and proteinase K were 0.28 and $0.63{\;}\mu\textrm{M}$, respectively, and for the cysteine proteases cathepsin B and papain were 0.66 and $0.28{\;}\mu\textrm{M}$, respectively. These data indicate that MAPI is not a potent selective inhibitor of serine or cysteine proteases. Progress curves for the inhibition of three proteases by MAPI exhibithe characteristic patterns; MAPI exhibited slow-binding inhibition of cathepsin B. It was rapidly associated with chymotrypsin before the addition of substrate and then reactivation of MAPI-inhibited enzyme was investigated in the presence of substrate. On the other hand, MAPI-proteinase K interaction was typical for those classical inhibitors. When MAPI was incubated with trypsin, there was an extensive reduction in the ingibitory activities of MAPI corresponding to 66.5% inactivation of MAPI, indicating that trypsin-like protease may play a role in the decrease of the inhibitory activity during cultivation.

• Biomolecules & Therapeutics
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• 제4권4호
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• pp.385-390
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• 1996

General pharmacological properties of urinary trypsin inhibitor (UTI) following intravenous administration of 1,000,000 units/kg were examined in terms of effects on central nervous system, cardiovascular system, respiratory system, gastrointestinal system in mice, rats and rabbits. Administration of UTI (1,000,000 units/kg, iv) had no effect on central nervous system; no influences on pentobarbital sleeping time, spontaneous activity, normal body temperature, chemoshock produced by pentylenetetrazole solution, writhing syndromes induced by 0.6% acetic acid solution, and motor coordination of mice. The administration of UTI (1,000,000) units/kg, iv) in rats had no effect on systolic blood pressure and pulse rate. UTI (500,000 units/kg, iv) given to anesthetized rabbits showed no effect on respiratory rate. However, it showed significant elevation of respiratory rate at the concentration of 1,000,000 units/kg. Gastric secretion of rat and intestinal motility of mice were not influenced by the dose of 1,000,000 units/kg. In terms of autonomic nervous system, the material did not show direct effect and inhibitory or augmentative action of histamine- or acetylcholine-induced contraction at the concentration of 2,000 units/ml in the isolated ileum of guinea pig.

• Natural Product Sciences
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• 제18권3호
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• pp.141-146
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• 2012

Veronica peregrina (Scrophylariaceae) has been widely used as a Korean traditional medicine for the treatment of various pathological conditions including infection, hemorrhage and gastric ulcer. In the current study, we investigated the inhibitory effect of methanolic extracts of V. Peregrina (VPM) on the LPS-mediated nitric oxide (NO) production in mouse (C57BL/6) peritoneal macrophages. NO production was significantly down-regulated by the treatment of VPM dose dependently. To evaluate the mechanism of the inhibitory action of VPM on NO production, we performed iNOS enzyme activity assay and checked the change of inducible nitric oxide synthase (iNOS) levels by Western blotting. Although VPM did not affect iNOS enzyme activity, iNOS protein expression was attenuated by VPM indicating VPM inhibits NO production via suppression of iNOS enzyme expression. In addition, VPM attenuated the expression of another pro-inflammatory mediator such as cyclooxygenase-2 (COX-2) in a dose dependent manner. We also found that VPM can reduce trypsin-induced paw edema in mice. Based on this study, we suggest that V. peregrina may be beneficial in diseases which related to macrophage-mediated inflammatory disorders.