This document collects Center for Drug Evaluation and Research (CDER) and Center for Devices and Radiological Health (CDRH) guidance documents, citations to the primary literature, and other published and unpublished documents relevant to development and approval of drug/device combinations collected by the CDRH Division of Cardiovascular, Respiratory and Neurological Devices (DCRND). Since the Master Bibliography number represents an accession number, an alphabetical (by author) listing appears at the end of the document, Any citation marked with a, is on file in the DCRND offices, 340B, in the Piccard Building (HFZ-450), 1390 Piccard Avenue, Rockville, MD 20850.

To gain insight into the etiological mechanism of dementia and to develop clinically effective congnitive enhancers, it is required to prepare animal models with symptoms and mechanism resemble to that in human. Dementia is mainly classified into two types : senile type of Alzheimer's disease (STAD) and cerebral ischemia-induced one. As animal models of cerebral ischemia, a couple of types in rats have been introduced : one is the occlusion of bilateral carotid arteries-induced forebrain/global ischemia and the other is the occlusion of middle cerebral arteries-induced focal/regional ischemia.

There have reports suggested that cerebral blood flow (CBF) has decreased in patients with both senile dementia of the Alzheimer's type and multi-infarct dementia, which are characterized by marked cognitive impairments. In addition, recent studies have demonstrated that decrease of CBF precedes the onset of multi-infarct dementia. These findings further suggest that chronic reduction of CBF may play an important role in the formation and progression of cerebral vascular dementia. Although transient cerebral ischemia, based upon vascular “reperfusion”, is apparently not paralleling the clinical condition, the transient cerebral ischemia model is one of the major methods investigated and the other is the cerebral embolism operation. Cognitive impairment and neuronal damages have been fully studied using these transient and/or embolic ischemia models. There are, however, few investigations focused the attention on the influence of chronic decrease of CBF on cognitive processes. In the present study, we have chosen a chronic ischemic model which is produced by permanent occlusion of bilateral common carotid arteries (2VO) in rats to investigate the neuronal damage and cognitive deficits through radial maze performance. We investigated furtherly the effects of tetramethylpyrazine (TMP), a constituent isolated from Ligusticum Chuanxiong on such a model.

Alzheimer's disease is believed to be associated with the loss of cholinergic activity in the cortex and hippocampus. In addition, it has been reported that the monoaminergic systems which also controls brain functions are disturbed in Alzheimer's patients. Based on these neurochemical background, a number of cholinesterase inhibitors including tacrine and its analogues and some monoamine oxidase inhibitors such as L-deprenyl and monoamine reuptake inhibitors have been developed for the treatment of dementia, but all of the known drugs are not truly effective. We thought that a drug that activates only one neurotransmitter system is not effective enough for the treatment of the symptoms associated with Alzheimer's disease and vascular dementia, and we conceived that an agent enhancing both central cholinergic and monoaminergic functions would be useful for the treatment of dementia

황금 뿌리로부터 Phospholipase-A2 저해제의 분리, 구조결정 방법: 황금의 MeOH 추출물로부터 용매분획과 column chromatography를 통하여 Phospholipase A2 저해작용을 가진 물질들을 분리하고, 이들의 구조를 확인하였다. 결과: Bioassay를 기초로 하여 Phospholipase-A2 저해작용이 있는 기지물질 6종을 분리, 구조확인 하였다.

자연계에 널리 분포하고 있는 flavonoid 유도체들은 다양한 구조에 따라, 다양한 생물활성을 보인다. 최근에는 이들이 cyclooxygenase/lipoxygenase를 저해한다는 사실이 알려진 후에 이들 유도체들을 항염증제 또는 항알레르기 약물로 개발하려는 시도가 계속되고 있다. 이들 연구의 일환으로 본 연구자들은 여러 flavonoid유도체들에 대한 in vivo 항염증능 및 림프구증식억제능을 보고하였다 (Arch. Pharm. Res. 16, 18, 25 (1993), 17, 31, 236 (1994), Life Sci. 54, 313 (1994)). 또한 flavonoid 유도체들 중 apigenin-dimer인 수종의 biflavonoid들이 group II phospholipase $A_2$ 저해제인 것을 밝혔고 (BBRC 205, 843 (1994)), 이들 중 몇 종은 림프구증식능에 대하여 억제작용을 보인다는 것을 알았다 (Life Sci. in revision). 본 연구에서는 이들 biflavonoid 유도체들 중 amentoflavone과 ginkgetin을 중심으로 이들의 림프구증식억제능 및 in vivo에서의 항염증작용을 연구하였다. 그 결과, ginkgetin (1-10 $\mu$M)은 Con A와 LPS에 의해 유도되는 림프구중식을 비가역적으로 억제하였으며, 이들 유도체들은 마우스 귀부종에 대하여 복강내투여에서 강력한 항염증능을 보인다는 것을 알았다.

1) 효소원의 조제 : 류마티스 관절염환자의 관절액 및 rat platelet PLA$_2$는 장등의 방법으로 조재하여 사용하였으며, 기질은 1-pal- (1-$^{14}$ C) linoleoyl PE로 하여 Dole 변법으로 효소활성을 측정하였다. 2) Histamine 유리반응 ; Rat복강으로부터 정제한 비만세포를 항원-항체 복합체로 자극하거나, $A_{23187}$등의 처리로 유리되는 histamine 을 methylation 시킨 후 유기용매법으로 추출한 후 scintillation counter로 측정하였다. 결과 : \circled1 천연물로부터 분리한 5종의 biflavonoid (amentoflavone 및 그 유도체)의 PLA$_2$ 저해 활성을 검토한 결과 거의 유사한 IC50 (약 3$\mu\textrm{m}$)을 나타내었다. \circled2 이들 중 amentoflavone은 염증성 PLA$_2$(Group II형 PLA$_2$)에 비교적 특이성을 나타내었다. 또한 제해양식은 비경쟁적 이면서 비가역적이였다. \circled3 비만세포에서 histamine 유리 억제반응은 자극제의 종류에 관계없이 억제작용을 나타내었으며, 기존에 임상적으로 사용되고 있는 Tranilast나 DSCG(disodium chromoglycate)에 비하여 10배 이상 강한 histamine 유리 억제작용을 나타내었다.다.

식물에서부터 새로운 소염작용제를 개발하기 위하여 여러식물을 대상으로 염증반응의 초기단계에서 중요한 역할을 하는 것으로 알려진 Phospholipase $A_2$에 저해활성을 갖는 물질을 검색하였고 그 중 강한 억제활성을 보인 유근피에서 유효성분을 분리하였다. 유근피의 에칠 아세테이트 분획에 대하여 실리카겔 크로마토그래피, Sephadex LH-20 크로마토그래피, 분취 박막 크로마토그래피를 수행하여 phenol성 -OH기를 갖는 활성성분인 PSH-II-84-1를 분리하였다. $^1$H-NMR 신호의 양상과 짝지움 상수 값에서 분리된 물질은 (+)-catechin 의 당 유도체로 확인되었다. $^{13}$C-NMR 자료를 분석하여 치환된 당은 D-apiofuranose로 확인되었다. 방향족환의 $^{l3}$C-NMR 신호들은 extended Huekel theory를 응용하여 얻은 net charge 계산 값과 상관시켜 할당하였다. 이상의 구조연구 결과 이 물질은 (+)-catechin-7-0-$\beta$-D-apiofuranoside로 밝혀졌다. (+)-catechin-7-0-$\beta$-D-apiofuranoside의 효소억제활성은 Type II Phospholipase $A_2$에 대하여 $IC_{50}$/이 600$\mu$M이었다.다.

후박의 주성분인 magnolol과 honokiol을 분리, 치주병인균 (혐기성그람음성균: Porphyromonas gingivalis 381, W50, A741-27, Prevotella intemedia 25611, 9336, G8-9K-3, 통기성그람음성균: Actinobacillus actinomycetemcomitans)에 대한 항균력을 조사하고 교원질 분해 활성, 섬유아세포재생능, 치은상피세포재생능, 치주인대세포재생능, cytokine생산 억제력시험을 하여 치주조직재생능을 관찰하였다. 후박과 대추의 안전성을 확인하기 위하여 일반임상증상, 체중측정, 피부반응을 관찰하였다. 아울러 이들 약물을 주성분으로 한 제제의 품질평가를 위하여 활성성분의 분리법 및 주성분인 magnolol과 honokiol을 지표로하여 HPLC 분석조건을 검토하였다.

새로운 혈소판 활성화인자 (PAF) 수용체 결합 저해활성 유효성분인 pinusolide ($IC_{50}$/=2.5$\times$$10^{-7}$M)와 arctigenin($IC_{50}$/=5.2$\times$$10^{-6}$M)을 측백엽(Biota orientalis)과 우방자(Arctium lappa)로부터 분리하여 이미 보고한 바 있다. PAF 수용체에 대한 구조활성 상관관계를 규명하고자 또한 용해도가 우수한 강력한 PAF 길항제를 개발하고자 이 길항제들의 유도체들을 다양한 유기화학반응을 이용하여 합성하였고 in vitro PAF receptor binding assay로 그 활성의 강도를 비교 검토하였다. 그 결과, lactone ring 부분은 unsaturated lactone이 saturated lactone보다 활성이 강하였고 ring이 개열되면 활성이 현저히 감소하였다. pinusolide 경우 exocylclic double bond 부분은 sp$^2$구조의 hydrophobic한 유도체가 활성이 강하였으며, methyl ester 부분은 hydrophobic한 유도체가 활성이 강하였다. Arctigenin의 경우 aromatic unit가 hydrophobic할수록 활성이 강하였다.

1993년 국내토양으로부터 분리동정된 Streptomyces속 균주중 항종양성이 우수한 균주 3주(DKM104, DKM117, DKM409)를 선정하고 이들을 대량 배양하여 종양억제인자를 분리하고 시험관내 종양세포 독성능 및 생체내 항암활성능과 이들 물질생산과 PLASMID DNA와의 관련성, $LD_{50}$등을 시험하여 새로운 항종양물질의 개발을 목적으로 하였다. 분리선별균주의 배양조건을 확립하였으며 배양여액을 국성이 적은 유기용매로부터 큰쪽으로 단계적으로 유효성분을 추출하여 Gel Chromatography를 이용하여 유효성분을 분획하였다. 시험관내 항종양능 시험은 MTT colorimetric검정법을 이용하여 $IC_{50}$값을 산정하였으며 생체내 항종양능은 마국의 NIC에서 권장하는 tumor panel system에 따랐다. 선정된 균주의 plasmid 분리는 alkalin lysis법을 채택하였으며 agarose gel electrophoresis로 plasmid profile을 시험하였다. Novobiocin등을 이용하여 Curing test를 시행하였고 독성실험은 $LD_{50}$량을 구해 항암효과 측정시에 Maximum dost로 투여하였고 최고용량을 기준으로 일정한 공비를 적응하여 3단계의 투여량을 설정하였다.(중략)

한국산 천연자원중 한방이나 민간요법에서 항종양제로 빈번히 사용되어온 생약들 중에서 103종을 선정하여 이들 성분들을 추출하고 시험관내에서 항종양성이 우수하고 정상세포에 손상을 적게 주는 생약 6종을 선별하여 암세포주에 대한 독성능 (in vitro)과 항종양성 면역감시기구(in vivo)및 LD$_{50}$등을 측정하여 항종양제로의 신약개발을 목적으로 수행하였다. 방법: 선별된 6종의 생약유효성분을 METHANOL로 추출하여 조추출물을 얻었으며 이물질들을 순차적으로 각각의 유기용매로 추출, column chromatography법으로 분획하였으며 분획분에 대한 암세포독성능은 MTT colorimetric 검정법을 이용하여 IC$_{50}$값을 구하였다. 면역감시기구 측정방범으로는 Balb/c mouae암,수 각 10수씩에 P388암세포주를 접종한군과 접종하지 않은 실험군에 생약추출분획물 8.6mg/0.2ml씩 20일간 매일 경구 투여시키고 대조군에는 생리식염수 0.2ml씩을 매일 경구 투여시켜 NK cell의 활성 MIF Recombinant IL-2로 유도시킨 NK cell활성능, chemotaxis등을 측정하였다. 생체내 항종양능 시험은 tumor panel system에 따라 mouse leukemia cell을 사용하여 측정하였다. 각분획성분의 투여용량은 실험동물에서 독성실험결과로 LD$_{50}$량을 구해 항암효과 평가시에 Maximum dose로 하였고 최고용량을 기준으로 일정한 공비를 적응하여 3단계의 투여량을 설정하였다. (중략)

Farnesyl Protein Transferase (FPTase) catalyses a post-translational modification of Ras that is obligatory for the cell transforming activity of this oncogene protein. The screening of natural products to identify inhibitors of this enzyme as a potential anticancer agents, has led to the isolation of a novel lactone, from the hydroid Solanderia secunda. Solandelactone G has been isolated from the hydroid Solanderia secunda collected along the offshore of Jaejudo and Keomunde. The structure of the compound has been determined as cyclopropane containing C$\_$22/ fatty acid lactone on the basis of the combined spectral and chemical methods

Cholesteryl Ester Transfer Protein (CETP) mediates the transfer of cholesterol ester and triglyceride between high-density lipoprotein (HDL) and other low-density lipoproteins, therefore, it might influence HDL levels. The levels of HDL is closely related to the atherogenic diseases in human and there were several reports that the trasgenic mice expressing CETP had much worse atherosclerosis than non-expressing control one. Therefore, selective inhibitors of CETP have the potential to be used as antiatherosclerotic agents. Continued screening for potent inhibitors of CETP led to the isolation of Suberitenone B from marine sponge. Suberitenone B, sesterterpenoids of a new skeletal class have been isolated from the sponge Suberites sp. collected from King George Island the Antartic. The structure of the metabolite has been determined by NMR experiments and chemical methods.

Cholesteryl Ester Transfer Protein(CETP), a hydrophobic glycoprotein with molecular mass 74KDa, is a lipid transfer protein found in plasma which mediates the transfer of cholesterol ester and triglyceride between high-density lipoprotein (HDL) and other lipoproteins, therefore, it might influence HDL levels. The lipoprotein profile associated with human CETP deficiency (that are two Japanese families, high HDL and low LDL) has low atherogenic potential, raising the possibility that CETP inhibitors can be used as antiatherosclerotic drugs.

Mutated forms of the ras oncogenes are associated with about 30% of human tumors. The ras genes encode 21KDa proteins, called p21 or Ras, that are associated with the plasma membrane. FPTase is a dimeric enzyme that catalyses the transfer of the farnesyl group from farnesyl pyrophosphate onto cysteine 186 at the C-terminus of the Ras protein. This is mandatory process for triggering ras oncogene toward tumor formation. Therefore, selective inhibitors of FPTase have the potential to be used as antitumorgenic agents.

Acyl-CoA:cholesterol acyltransferase (ACAT, EC 2.3.1.26) plays an important role in the control of intracellular free cholesterol content via its cholesterol esterifying activity. ACAT inhibitors are expected to be effective for treatment of atherosclerosis and hypercholesterolemia. In the course of a screening program for ACAT inhibitors from microbial sources, GERI-BP001 M, A, and B were isolated from the fermentation broth of a fungal strain. GERI-BP001 compounds were isolated from a culture broth of Aspergillus fumigatus F37 by acetone extraction, EtOAc extraction, SiO$_2$ column chromatography, and reverse phase HPLC. The structure of GERI-BP001 coumpounds were determined by $^1$H-NMR, $\^$l3/C-NMR, 2D-NMR, NOESY, and long range C-H COSY experiments. GERI-BP001 M, A, and B inhibit ACAT activity in an enzyme assay system using rat liver microsomes by 50% at concentrations of 75, 147, and 71 ${\mu}$M, respectively.

청주 근처 토양에서 분리한 에너지 대사 저해제인 2,4-dinitrophenol (DNP)에 내성을 보이는 Streptomyces CM 001 균주가 생산하는 항진균성 물질(CUA)을 순수하게 분리 정제하고 화학분석을 통한 구조의 규명, 항진균활성 연구 및 유전적 변이원성(mutagenicity)등을 살펴보았다. CM 001 균주를 베네트 배지와 같이 영양성이 좋은 배지에서 배양시 항진균 활성이 동반 생성되는 색소와 비례적으로 증가했다. 배양 상등액 속의 항진균성 물질들을 염산처리, 유기용매처리 후 Amberlite XAD-4 column chromatography, silica gel adsorption-Sephadex LH-20 chromatography 과정을 통해 색소가 제거된 부분 정제 백색분말로 얻었다 이로부터 Prep. HPLC과정을 통하여 8종의 화합물을 각기 분리하였는데 이 화합물들은 동일한 UV spectrum과 216, 260nm에서 최대 흡수파장을 보였다.

The succinic semialdehyde dehydrogenase which is one of the key enzyme of GABA shunt in CNS has been purified from bovine brain homogeneously for the first time. The molecular mass of the native enzyme was estimated to be approximately 110,000 on gel filtration, The subunit molecular mass was determined by SDS-PAGE to be 54,000. These results indicate that the enzyme is a dimeric protein made up to identical subunits. Chemical modification studies of the enzyme suggest that the critical lysyl, connected with catalytic activity of the enzyme, The binding of IAF-SSDH(enzyme tagged with fluoreceine) to GABA transaminase which catalyzes the degradation of GABA was monitored by steady emission anisotropy. The changes of fluorescence anisotropy by interactions between two enzymes suggest that the formation of enzyme cluster must be invoved in the regulation of GABA concentration in brain tissues. The inhibitory effects of some antiepileptic and anticonvulsant drugs on the enzyme were also examined.

Monoclonal antibodies against bovine brain succinic semialdehyde reductase were produced and characterized. A total of nine monoclonal antibodies recognizing different epitopes of the enzyme were obtained, of which two inhibited the enzyme activity and three stained cytosol of rat spinal cord neurons as observed by indirect immunofluorescence microscopy. When unfractionated total proteins of bovine brain homogenate were seperated by gel electrophoresis and immunoblotted, the antibodies specifically recognized a single protein band of 34 kDa, which comigrates with purified bovine succinic semialdehyde reducatase Using the antisuccinic semialdehyde reductase antibodies as probes, we investigated the cross-reactivites of brain succinic semialdehyde reductases from some mammalian and an avian species. The immunoreactive bands on Western blots appeared to be the same in molecular mass-34 kDa-in all animal species tested, including humans. The result indicated that brain succinic semialdehyde reductase is distinct from other aldehyde reductases and that mammalian brains contain only one succinic semialdehyde reductase. Moreover, the enzymes among the species are imunologically very similar, although some properties of the enzymes reported previously were different from one another.

We investigated the effect of derivatized phospholipid, P-pyridoxyl dipalmiuylphosphatidylethanolamine (P-pyr-DPPE), on the catalytic activity of purified porcine brain glutamate decarboxylase(GAD) which catalyzes the synthesis of GABA known as major inhibitory neurotransmitter in CNS. When the P-pyr-DPPE was incorporated into dipalmitdylphosphatidylcholine(DPPC) or phosphatidylserine(PS) vesicles, these vesicles enhanced the catalytic activity of GAD. P-pyr-DPPE also interacted with apoglutamate decarboxylase(apoGAD) and produced the free pyridoxal-5-phosphate(PLP) which is the natural cofactor of GAD. This result indicated that apoGAD catalyzed the cleavage reaction of the P-pyridoxyl moiety of the derivatized phopholipid to generate free PLP, and then free PLP bound to the apoGAD resulting in restroration of the catalytic activity of the enzyme.

GABA transaminase is inactivated by preincubation with p$^1$, p$^2$-bis(5'-pyridoxal) diphosphate at pH 7.0. The inactivation under pseudo-first order conditions proceeds at a slow rate (K$\_$obs/=0.035 min$\^$-1/). The degree of labeling of the enzyme by p$^1$, p$^2$-bis(5'-pyridoxal) diphosphate was determined by absorption spectroscopy, The blocking of 2 lysyl residues/dimer is needed for inactivation of the transaminase. The time course of the reaction is significantly affected by the substrate ${\alpha}$-ketoglutarate, which afforded complete protection against the loss of the catalytic activity. Whereas cofator pyridoxal phosphate failed to prevent the inactivation of the enzyme. Therefore, it is postulated that binding of ${\alpha}$-ketoglutarate tn lysyl residues is the major factor contributing to stabilization of the catalytic site and bifuctional reagent p$^1$, p$^2$bis(5'-pyridoxal) diphosphate blocks lysyl residues other than those involved in the binding of the cofactor.

Succinic semialdehyde reductase, one of key enzyme of GABA shunt in CNS, is inactivated by o-phthalaldehyde, The inactivation followed pseudo first-order kinetics, and the second-order rate constant for the inactivation process was 28 M$\^$-1/s$\^$-1/ at pH 7.4 and 25$^{\circ}C$. The absorption spectrum(λ$\_$max/=377nm), fluorescence exitation(λ$\_$max/=340nm) and fluorescence emission spectra (λ$\_$max/=409nm) were consistent with the formation of an isoindole derivative in the catalytic site between a cysteine and a lysine residues about 3${\AA}$ apart. The substrate, succinic semialdehyde, did not protect the enzymatic activity against inactivation, whereas the coenzyme, NADPH, protected against o-phthalaldehyde induced inactivation of the enzyme. About 1 isoindole group per moi of the enzyme was formed following complete loss of the enzymatic activity. These results suggest that the amino acid residues of the enzyme participating in reaction with o-phthalaldehyde more likely residues at or near the coenzyme binding site.

Virus-encoded HIV-1 reverse transcriptase (RTase) is one of the major targets for the development of drugs for HIV-1 since it is an essential enzyme-for the replication cycle of HIV-1. We cloned the entire reverse trancriptase gene into an inducible expression vector with tac promotor= RTase was stably overexpressed and induced by IPTG and the highly-expressed RTase was purified partially by use of DEAE cellulose and Mono Q column. The partially purified enzyme (663kDa, 51kDa) as exhibited by SDS-PAGE showed the high specific activity (16,570U/mg) when the assay for the RTase activity was carried out using $^3$H-dTTP and poly(rA): oligo(dT)12-18 as the substrate.

Human immunodeficiency virus type 1 (HIV-1) contains several nonstructural genes which are required for the viral replication and disease pathogenesis. Among them, tat and nef genes encode an essential transactivator of HIV-1 LTR and a pluripotent protein which seems to be essential for the in vivo but not in vitro viral replication, respectively. We constructed two tat and n of defective HIV-1 and tested for their ability to replicate in several T cells. The defective viruses did not replicate in CD4$\^$+/ T cells, but rescued in the recombinant Jurkat-tat cell which also contains tat gene. The replication of tat and nef defective HIV-1 which expresses chloramphenicol acetyltransferase(CAT) gene was easily detected by a sensitive CAT assay. No revertant was identified during the passages of the mutant viruses for more than two months in Jurkat-tat cells. tat and n of defective HIV-1 could be used instead of wild type viruse for several purposes such as inhibitor screening and development of attenuated AIDS vaccine.

Increasing evidence indicates that the production of nitric oxide (NO) by inducible NO synthase (NOS) is tightely regulated. Transforming growth factor-${\beta}$ (TGF-${\beta}$) is a homodimeric protein secreted during macrophage activation, but several lines of evidence suggest that TGF-${\beta}$ is selectively suppressive for macrophage NO production. We therefore reasoned that a strategy employing oligodeoxynucleotides(ODNs) complemently to TGF-${\beta}$ mRNA (antisense ODNs) might increase NO production in IFN-${\gamma}$-treated murine peritoneal macrophages. To evaluate this concept, we tested the effects of antisense ODNs targeted to TGF-${\beta}$ mRNA (25-mer ODNs complemently to TGF-${\beta}$mRNA sequences) by introducing it into the medium of cultured macrophages. Phosphorothiolation of ODNs were employed to retard their degradation. Antisense ODNs had no effect on NO production by itself, whereas IFN-${\gamma}$ alone had modest effect. When antisense ODNs were used in combination with IFN-${\gamma}$, there was a marked cooperative induction of NO production, These effects of antisense ODNs were associated with decreased TGF-${\beta}$ expression in activated macrophages. ODNs with the same nucleotides but a scrambled sequence had no effect. Adding anti-TGF-${\beta}$ antibodies to the IFN-${\gamma}$-treated macrophages mimicked the positive effect of antisense ODNs on NO production. In addition, the effects of either antisense ODNs or anti-TGF-${\beta}$ antibodies were blocked by adding TGF-${\beta}$ in cultured macrophages. These results indicate that the generation of TGF-${\beta}$ by activated macrophages provides a self-regulating mechanism by which the temporal and perhaps spatial production of NO, a reactive and potentially toxic mediator, can be finely regulated.

Although synthetic antisense oligodeoxynucleotides (ODNs) have been used to dissect gene function in vitro, technical difficulties of targeted delivery prevented the use of this approach for investigating the effect of gene products in vivo. Here we report the use of local delivery of antisense transforming growth factor-${\beta}$l (TGF-${\beta}$1) oligonucleotides to decrease the fibrosis in the skin wound. Adult wounds heal with scar-tissue formation, whereas fetal wounds heal without scarring and with a lesser inflammatory and cytokine response. We reasoned that strategy emptying antisense TGF-${\beta}$1 ODNs complementary to TGF-${\beta}$1 mRNA might decrease the scarring in dermal wound of mouse. To evaluate this concept, we tested the effects of antisense ODNs targeted to TGF-${\beta}$1 mRNA by topical application of the chemical on the skin wound. Phosphorothioate antisense ODNs was employed to retard their degradation. When antisense TGF-${\beta}$1 ODNs were applied into the wound site, there was a maked reduction of scar compared with control wound site, These effects of antisense TGF-${\beta}$1 ODNs on the scar formation were associated with decreased expression of TGF-${\beta}$1 gene. However sense TGF-${\beta}$l ODNs had no effect on expression of TGF-${\beta}$1 gene. Also, control wounds healed with excessive fibrosis, whereas the antisense treated wounds healed with less fibrosis. In conclusion, our results indicate that antisense TGF-${\beta}$1 ODNs could be used for amelioating scar formation during wound healing.

T4 Endonuclease V (Mw 16,000) acts as a repair enzyme for UV induced pyrimidine dimers in DNA. Many researchers have studied the biochemical characteristics of the enzyme. However the precise action mechanism of T4 endo V has not fully elucidated yet. In our laboratory NMR spectroscopy technique is being used for the structural study of T4 endo V. Because of its low temperature stability and high content of ${\alpha}$-helix, the conventional $^1$H NMR technique was inapplicable. Therefore we utilized stable isotope labeling technique and so far prepared about 10 amino acid specific labeled proteins. The HSQC spectra of amino acid specific labeled proteins will help us to interpret the triple resonance 3D, 4D data which are under processing, We also studied the behaviors of specific amino acid residues whose roles might be critical. When the enzyme labeled by $\^$15/N-Thr was mixed with the substrate oligonucleotide (semispecific -TT- sequence), one crosspeak in its HSQC spectrum was completely desappeared, which means that one of seven Thr residues is in the binding site of the enzyme with DNA, This result is well consistent with previous report that implicated the Thr 2 residue in the activity of the enzyme. Similar studies were carried on the behaviors of Arg and Tyr residues.

It is well known that Src Homology 2(SH2) domain in many intracellular signal transduction proteins is very important. The domain has about 100 amino acid residues and bind phosphotyrosine-containing peptide with high affinity and specificity. Lck SH2 domain is a Src-like, lymphocyte-specific tyrosine kinase. An 11-residue phosphopeptide derived from the hamster polvoma middle-T antigen, EPQp YEEIPIYL, binds with an 1 nM dissociation constant to Lck SH2 domain. And it is known that the phosphotyrosine and isoleucine residues of the peptide are tightly bound by two well-defined pockets on Lck SH2 domain's surface. To investigate the conformational changes during complexation of SH2 domain with phosphopeptide we have performed the molecular dynamics simulation for Lck SH2 domain with peptide and peptide-free form at look in aqueous solution. More than 3000 water molecules were incorporated to solvate Lck SH2 domain and peptide. Periodic boundary condition has been applied in molecular dynamics simulation. Data analysis with the results of that simulation shows that the phosphopeptide makes primary interaction with the Lck SH2 domain at six central residues, The comparison of the complexed and uncomplexed SH2 domain structures in solution has revealed only relatively small change. But the hydrophilic and hydrophobic pockets in the protein surface show the conformational changes in spite of the small structural difference between the complex and peptide-free forms.

4-lBB molecule is expressed on the surface of activated CD4$\^$+/ and CD8$\^$+/ T cells. We generated a panel of anti-4-1 B5 murine mAbs using a fusion protein consisting of the extracellular domain of human 4-1 BB fused to Glutathione S-transferase. The binding activity against cell surface 4-1 BB molecule was assessed by flow cytometry analysis. These studies showed that several anti-4-1 BB mAbs bound to 10-30% of CD4$\^$+/ and CD8$\^$+/T cells in PHA or Con A stimulated PBLs, although these mAbs interacted with only, l-2% of CD4$\^$+/ and CD8$\^$+/ T cells in normal PBLs, indicating the specificity of mAbs to the 4-l BB molecule on activated CD4$\^$+/ and CD8$\^$+/ T cells. Next, we examined the effect of an anti-4-l BB mAb (4B4-1-1) on allogeneic mixed lymphocyte reactions (MLRs). The data indicated that the antibody significantly inhibited the proliferative response at higher concentrations. When tested with several T cell mitogens, the antibody had no stimulatory or inhibitory effects on the mitogen-mediated T cell proliferation. These data suggest that 4-1 BB molecule may play a role in the regulation of antigen-mediated immune response.

전년도에 ara-C 유도체 중에서 ara-CDP-DL-PTBA가 각종 암세포주에 대한 뛰어난 항암작용이 보고된바 있으며, 이후 시료의 대량확보, Bulk Formulation 및 전임상등의 세가지 분야에 관해 연구를 수행하였다. Ara-CDP-DL-PTBA를 수용액에 현탁, 초음파분쇄후 NICOMP Analysis에 의하여 micellar solution의 입자도를 알아 본 결과 fresh prepared micelles은 11,1mm size가 88.63%이며 50,5mm가 11.37%로 나타나 평균 19,0mm가 되고, Reconstructed micells은 10.9mm size가 99.87%이며 356.1mm가 0.13%로 나타나 평균 11.0mm가 된다. Ara-C와 Ara-CDP-DL-PTBA의 대사작용을 알아본 결과, ara-C는 투여 1시간째 2,850 $\pm$ 450 pmol, 4시간째 450$\pm$190 pmol, 24시간째 30 pmol 이하로 ara-CTP의 혈중 농도가 급격히 감소하는 반면에 ara-CDP-DL-PTBA는 1시간째 650$\pm$120 pmol, 2시간째 1,800$\pm$500 pmol, 24시간째 300$\pm$90 pmol으로 ara-CTP의 혈중 농도가 서서히 감소하였다.

The analysis of membrane receptors for hormones and neurotransmitters has progressed considerably by pharmacological and biochemical means and more recently by the use of specific anti-receptor antibodies. A 14-mer peptide (from Phe102 to Leu115 of ${\beta}$2-adrenergic receptor) was synthesized and this peptide was coupled to carrier protein Keyhole Limpet Hemocyanin(KLH) by glutaraldehyde method. A 0.5mg of KLH-coupled peptide was emulsified with equal volume of complete Freund's adjuvant and injected via popliteal lymph node to each of the three Newzealnd White rabbits. Booster injections were repeated at 4 weeks interval for three times with incomplete Freund's adjuvants. One week after the final injection, serum was prepared from ear artery. Nonspecific immunoglobulins were removed by passing the serum through KLH-Sepharose 6B affinity matrix and further by incubation with bovine lung aceton powder. The titer of the antibody for synthetic peptide which was determined by enzyme linked immunosorbent assay(ELISA) was about l/l,000. The antibody produced in this study revealed 67kDa protein band in the western blot of partially purified guinea pig lung ${\beta}$-adrenergic receptor preparation. The antibody inhibited ${\beta}$-adrenergic antaginist [3H] Dihydroalprenolol binding to soluble ${\beta}$-adrenergic receptor by 25% while control sera did not show any inhibitory effects, The result of this study suggests that the peptide sequence selected in this study may play some important roles in adrenergic receptor-ligand interaction.

The muscarinic effects of carbachol were compared on the isolated ileums of guinea-pig, rat and rabbit to elucidate the underlying mechanism of species differences in sensitivity for carbachol. The ED$\_$50/ value estimated on the guinea-pig ileum was 4 to 6-fold lower than those obtained on the rat and rabbit ileums, but the K$\_$A/ values of carbachol determined by functional assays were almost identical with 12-l7 ${\mu}$M in all of three ileums. The competition data of carbachol for [$^3$H]QNB binding were best described by a two-site model yielding the Ki values of 0.4-0.6${\mu}$M and 12-16${\mu}$M for high(K$\_$H/) and low(K$\_$L/) affinity sites, respectively. The low affinity dissociation constants(K$\_$L/) of carbachol determined from receptor binding studies thus were not significantly different from the K$\_$A/ values estimated from functional studies. The percentage of receptor occupation that carbachol requires for half-maximal response was approximately 3 to 5-fold lower in guinea-pig compared to rat and rabbit whereas the density of muscarinic binding sites per gram of ileum measured by [$^3$H]QNB saturation isotherms was two-fold higher in guinea-pig than that in rat and rabbit. Therefore, the numbers of muscarinic receptors occupied at ED$\_$50/ values of carbachol were about two-fold lower in guinea-pig, suggesting two-fold greater intrinsic efficacy. These results indicate that the guinea-pig ileum has higher muscarinic receptor density and greater intrinsic efficacy for carbachol than the rat and rabbit ileums.

Tetrahydroisoquinoline (THI) compounds have various pharmacological actions in the cardiovascular system. Recently, we have synthesized 1-${\alpha}$-naphthylmethyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, YS 49. In the present study, we evaluated the effect of YS-49 on positive inotropic and chronotropic action using isolated rat heart and on blood pressure and heart rate using anesthesized rabbit. Vasodilating action was also assessed in isolated rat thoracic aorta. YS 49, concentration-dependently relaxed rat aorta precontracted with phenylephrine (PE, 0.3 ${\mu}$M) and high potassium (high K$\^$+/, 65.4 mM). The 50% inhibitory concentration (IC$\sub$50/) of YS 49 in PE-induced and high K$\^$+/-induded contraction was 5.36 ${\mu}$M and 2.52 ${\mu}$M, respectively. In isolated rat atria, YS 49 increased both heart rate and force, and in anesthesized rabbit it decreased blood pressure but increased heart rate. In addition, to know the mechanism of action of the compound, propranolol, nonselective ${\beta}$-antagonist, and phentolamine, ${\alpha}$-blocker, were used. Furthermore, a comparison with the effect of higenamine, trimetoquinol on the vasodilating action in rat thoracic aorta was also made. The action of YS 49 was inhibited by the presence of propranolo, not pentolamine. These results indicate that cardiotonic and vasodilatory action of YS 49 is attributable, at least in part, for ${\beta}$-receptor stimulation.

Surface epithelial cells isolated from hamster tracheas and grown on a thick collagen gel become a highly enriched population of mucus-secreting cells. Epithelial cells from tracheas of hamsters were collected using enzymatic procedures and cultured under various conditions. The medium used consisted of a 1:1 mixture of medium 199 and Dulbecco's modified Eagle's (DME) medium which was conditioned before use. Insulin, transferrin, hydrocortisone, epidermal growth factor, and extract from bovine hypothalamus were used as supplement. Due to relatively low basal rates of min secretion from in vitro cultures, cultures are generally radiolabeled using $^3$H-glucosamine as a metabolic precursor. The radiolabeled mucinsreleased are quantitated by precipitation with TCA/PTA. Using this cell culture system, we investigated mucin release of goblet cells by altering the media bathing the apical surface of hamster tracheal surface epithelial(HTSE) cells. Acidic media added sulfuric acid caused sigcificant increases in mucin relesse (155${\pm}$20% at pH 4 and 146${\pm}$16% at, pH 5). Ammonium hydroxide also increased mucin release at pH 9.0(156${\pm}$17%) and pH 10(295${\pm}$9%) respectively. This additional mucin release seems to be associated with cell membrane damage as indicated by release of cellular LDH. SP stimulates secretion of mucin in cultured HTSE cells(154${\pm}$16% at 1${\times}$10$\^$-6/M and 165${\pm}$25% at 1${\times}$10$\^$-5/M. PAF at 5${\times}$10$\^$-6/M and 5${\times}$10$\^$-5/M enhanced by HTSE cells in vitro 168${\pm}$34% and 259${\pm}$30% of mucin secretion, respectively. The increase in mucin release by PAF and SP was not secondary to cell damage or necrosis. SP and PAF may be in mediating mucous secretion induced by inflammation irritantion and infection.

The Influence of age on the endothelial modulation of angiotensin II (AII)-induced contractile response was investigated in isolated aortic rings of rats ranging in age from 0.7 to 20 months. Hemoglobin and L-NAME were used to examine whether age-related changes in the EDRF-releasing system were involved in endothelial modulation of All-induced contraction in rat aorta. In all five age groups (0.7, 1.5, 3, 6, 20 months), hemoglobin (10 ${\mu}$M) significantly enhanced All-induced contractile response only in aorta with endothelium intact. L-NAME (10 ${\mu}$M) Produced a significant enhancement in All responses in endothelium-intact aortas from rats aged 0.7 and 1.5 months, but it had no effect in aortas from older rats aged 6 and 20 months. Indomethacin (10 ${\mu}$M) did not affect All-induced contractile responses in both endothelium intact and removed aortas from rats at the age of 0.7 to 20 months. Hemoglobin (10 ${\mu}$M) abolished acetylcholine-induced relaxation response in aortas from young and old rats. L-NAME completely abolished the relaxation in aortas from young (0.7 and 1.5 months), but incompletely in aortas from older (6 and 20 months) rats. The sensitivity of endothelium-dependent relaxation to A23187 increased with age between ages of 0.7 and 6 months, with no further increase noted up to 20 months of age. These results suggest that endothelial modulation of AII-induced contraction in rat aorta might involve age-related alteration in EDRF-releasing system, probably via post-receptor mechanism.

에탄올에 의한 급성독성은 에탄올, 아세트알데히드 및 에탄을 대사산물의 변형생성물질 등에 기인하므로 알콜 섭취 후 혈중 에탄을 농도 및 아세트알데히드의 농도를 낮추는 것은 음주에 의한 급성 중독상태에 머무는 시간을 단축시키는데 중요하다. 본 실험에서는 Bacillus subtilis natto sp.를 식물 추출액을 배지로 하여 배양한 후 단백 분해효소로 고분자물질을 절단하여 얻은 발효액인 바이오짐(상품명: Biozyme)을 주성분으로 한 '비지니스(조선무약합자회사)' 의 인체 혈중 알콜대사 촉진작용을 검색하였으며, 비지니스 및 바이오짐이 랫드의 에탄을 대사에 미치는 효과를 검토하여 다음과 같은 결과를 얻었다. 인체 혈중 알콜대사 촉진작용을 검색하기 위해 알콜 섭취 전과 후의 혈중 에탄올 농도를 비교하였는데, 대조군에 비해 비지니스 투여군이 30분 후부터 2시간 후까지 혈청 알콜농도가 낮게 나타났다. 대조군의 혈청 알콜농도를 100으로 하였을 때 알콜 투여 30분, 60분, 90분, 120분 경과 후 시험군은 각각 대조군의 84.3%, 89,0%, 85.9%, 75.8%를 나타내어 평균 16% 정도 낮았다. 시험군의 AUC는 대조군의 AUC의 89%로 비지니스 투여군에서 혈액내 알콜의 제거가 빠르게 진전된다는 것을 보여주었다. 또한, 랫드의 에탄을 대사에 미치는 효과를 알아보고자 바이오짐 및 비지니스 투여 후 채혈하여 혈중 에탄을 및 아세트알데히드 농도를 측정하였다. 비즈니스 투여시 혈중 알콜 농도는 알콜 투여 60분 경과후 가장 큰 감소 효과(대조군:83.70$\pm$11.80mg/이, 시험군:45.12$\pm$6.63mg/d1, 47% 감소)를 나타내었으며, 시험군의 AUC는 대조군에 비해 30% 감소하였다. 혈중 아세트알데히드 농도는 투여 60분 후 비지니스 투여군(4.56$\pm$0.51nmol/$m\ell$)이 대조군(6.45$\pm$0.64nmo1/$m\ell$)에 비해 유의성 있는 감소(29%)를 나타내었으며, 시험군의 AUC는 대조군에 비해 35% 감소하였다. 바이오짐 투여시 혈중 에탄을 농도가 알콜 투여 2시간 경과 후 가장 큰 감소 효과(대조군:49.10$\pm$5.20mg/dl, 시험군:25.90$\pm$7.16mg/d1, 47% 감소)를 나타내었으며, 시험군의 AUC는 대조군에 비해 39% 감소하였고, 혈중 아세트알데히드의 농도는 투여 60분후 시험군(3.96$\pm$0.07nmo1/$m\ell$)이 대조군(6.45$\pm$0,64nmo1/$m\ell$)에 비해 유의성 있는 감소(39%)를 나타내었으며, 시험군의 AUC는 대조군에 비해 48% 감소하였다 한편, 시험관내 에탄올 대사 효소에 대한 바이오짐의 효과를 검색해본 결과 바이오짐(2.0 $\mu\textrm{g}$/assay)에 의해 Aldehyde dehydrogenase(1.5unit/assay)의 활성이 14% 증가되었다. 본 연구의 결과로 볼 때, 비지니스 및 바이오짐은 음주 후 상승된 혈중 에탄을 농도 및 아세트알데히드의 농도를 현저히 감소시키는 효과가 있었다.

It has been suggested that oxygen-derived free radicals have an important role in the pathophysiology of acute gastric ulceration induced by NSAIDs and ischemia-reperfusion. Taurine is hypothetized to exert its protective effect on NSAIDS-induced gastric injury by its antioxidant properties, Protect ive effect of taurine on indomethacin-induced gastric mucosal lesion and its protective mechanism were investigated. Intragastric administration of 25 mg/kg of indomethacin induced hemorrhagic lesions on the glandular stomach in rats, Pretreatment with 0.25 g/kg of taurine for 3 days significantly reduced the gastric lesion formation and Inhibited the elevation of lipid peroxide level In gastric mucosa. Both resting and FMLP-induced luminol-dependent chemiluminescence of rat peritoneal neutrophils increased immediately after treatment of indomethacin. 5-20mM of taurine inhibited chemiluminescence of neutrophils activated by indomethacin and/or FMLP. Human neutrophils (polymorphonuclear leukocytes) significantly adhered to confluent monolayer of human umbilical vein endothelial cells(HUVEC) after coincubation with aspirin or indomethacin. Also taurine prevented neutrophil adhesion induced by these drugs to HUVEC in dose-dependent manner. These results indicate that the protective effect of taurine against NSAIDS-induced gastric mucosal Injury is due to its antioxidant effect, which inhibits lipid peroxidation and neutrophil activation.

The three combined products were prepared as suspended solutions composed of various ratio of Sucralfate, Hydrotalcite and Neusilin, into which 30% ethanol extracts of Machili cortex, and of Atractylodis rhizoma were added. The efficacy for these products was examined in vivo using a pyrous ligation method in rats. The influence of these products on the intestinal motility was also examined in mice. In all experimental setting, the antisecretory effect of the combined treatment was more pronounced than that of each drug alone. The combined treatment consisted of Sucralfate, Hydrotalcite, Neusilin ratios of 2:2:1 produced the highest inhibitory effect for the gastric secretion. The intestinal motility was not influenced significantly by the treatment of all experimental setting. The above results revealed that the therapeutic dose of Sucralfate, Hydrotalcite, Neusilin given in combination showed a synergistic effect for the inhibition of gastric secretion and little side effect on the intestinal motility. Therefore, the combined product with Sucralfate, Hydrotalcite, Neusilin ratio of 2:2:1 is recommended for the useful drug to heal the gastrointestinal diseases with no side effect on the intestinal motility.

Aspalatone (APT)의 예상되는 항산화 작용을 검색하기 위하여 Doxorubicin(Dox.) 유발독성 (Dox.의 작용 발현 기전은 oxyradical 생성에 의해 매개됨)으로 인하여 변화되는 항산화 효소계 및 과산화지질 생성에 대하여 APT의 효과를 검색하고, 별도로 Hydroxyl radical 생성에 미치는 APT의 효능을 검정하기 위하여 ESR spin trapping technique을 이용하여 ㆍOH/DMPO ESR signal을 검정하여 아래의 결과를 얻었다. Dox 투여 2일 후에 생쥐 심장에서 얻어진 SOD-1의 보상성 유도는 APT 병용 투여로 억제되었고, Catalase 활성을 유도하였으나 MBA 치에는 유의한 변화가 나타나지 않았다. 반면에 간에서는 APT으로 인하여 GSH-PX가 현저히 유도되었고 (다른 항산화 효소에는 유의한 변화가 나타나지 않았음), MDA 치가 억제되었다. 한편 Fenton 반응동안 증가되었던 ㆍOH/DMPO ESR signal은 APT에 의해 억제되었다. 이상의 결과에서 APT의 항산화성 효능이 인정되지만, Dox. 및 APT 각각의 용량 및 노출 시간의 상관성, 그리고 생화학적인 결과에 대한 (미세)조직학적인 확인이 요구되며 각각의 실험군, 즉 aspirin, maltol에 대한 효능/효력 검정도 추가로 보완되어야 할 것으로 생각된다.

뇌신경 변성 / 퇴행과 관련된 중요한 병인론 중의 하나는 변성 과정에서 형성된 유리기(free radical)로 인한 항산화계의 평형 소실로 알려져 있다. Aspalatone (APT)의 예상되는 항산화 효능을 검정하기 위하여 본 실험에서는 Kainic acid (KA) 유발 뇌변성 모델을 적용하였다. KA 모델은 변연계의 간질성 경련과 신경세포 변성에 대하여 재현성 있는 병변 모델을 제공해 주며, 이와 같은 신경세포의 병독 기전에 산소 유리기가 관여함이 강력히 시사되고 있기 때문이다. KA 투여로 인하여 지속적이고도 전형적인 간질성 경련이 관찰되고 1일 이내에 높은 치사율을 보였으나 APT으로 인하여 그 간질성 경련 행위와 비율이 억제되고 KA 유발 치사율도 억제되었다. 최종 KA 투여 3일 후에 얻어진 흰쥐 해마 및 대뇌 피질에서 항산화 효소인 Superoxide dismutase (SOD), Catalase (Cat.), Glutathione peroxidase (GSH-PX) 및 과산화지질의 지표인 Malondialdehyde (MDA)를 검정하였다. 대조군에 비하여 KA는 뇌조직의 SOD-1을 유도하였으나, Cat.와 GSH-PX의 활성은 현저히 유도되지 않았고, 반면에 MDA 치는 현저히 증가하였다. 즉, Cat., GSH-PX와 같은 $H_2O$$_2$중화제가 동반 유도하지 않는 SOD의 유도는 세포내 축적되는 $H_2O$$_2$로 인하여 Fenton/Haber-Weiss 반응을 가속화하여 과산화지질화를 촉진함을 시사한다. APT 병용 투여로 SOD는 현저히 유도되지 않았으나 특히 Cat.가 현저히 유도되어지고 MDA는 억제되었다. 이와 같은 생화학적인 결과는 다음의 형태학적인 소견과 일치한다. Fos 관련 항원 (FRA)와 SOD-1을 면역세포화학 (Immunocytochemistry)적 방법으로 이중 표식 (double-labelling) 하였다. FRA는 KA로 인한 신경세포의 자극에 대한 지표로 응용하였고, SOD-1은 퇴행성 뇌질환에서 산화적 손상의 지표로 사용하였다. KA 투여로 해마의 dentate gyrus (DG) 내에 강한 면역환성 (immunoreactivity)이 나타났고 pyramidal cell layer (PCL)와 glia에 SOD-1이 강하게 염색되었다. APT 병용 투여로 상당수의 경련이 일어나지 않은 흰쥐는 해마의 DG에 FRA가 경미하게 염색되었고, PCL에 SOD-1도 경미하게 나타났으나, 경련이 나타난 쥐에서는 KA만을 투여한 흰쥐와 구별되지 않았다. 이상의 APT의 항산화 효과는 KA로 인한 뇌세포 변성 개선에 중요한 인자로 작용할 것으로 사료되나, 보다 명확한 APT의 기전을 검색하고 직접 임상에 응응하기 위하여는 보다 다양한 실험 조건이 보완되어야 찰 것으로 생각된다.

It was recognized that lead intoxication reduces thiamine content in the brain of rat and this change produces the alterations of thiamine-related biochemical reactions. In the present study, it was tested whether the changes of myelin composition as well as seizure threshold induced by lead intoxication in rats may be related to these changes of thiamine status and thiamine related biochemical factors. Wistar rats were divided into five groups: Control group, Lead-treated group, Lead plus Thiamine-treated group, Thiamine-deficient group, Pyrithiamine-treated group. Each group was divided into three subgroups: 3, 7 and 16 week old group. Myelin protein and phospholipid, one of the compositions of myelin lipid, were measured in the myelin isolated from rat brain. Threshold of electric shock seizure was tested in each group. The amount of each myelin composition in lead-treated group and thiamine-deficient group was significantly lower than those of all the brains in control group, but recovery by supplementation with thiamine during lead intoxication was occurred only in the cerebellum of 3 week old animal. Thresholds of the electric shock seizure of lead treated group and thiamine deficient group in 3 and 7 week old rats were significantly lower than those of control group, while those of lead plus thiamine treated group were similar to those of control group.

On the combination of antacid, the pharmacokinetics and gastric adhesion of $\^$14/C-aceglutamide aluminum complex($\^$14/C-AGA) were examined in rats. Specially, This study was focused on the drug interaction that the coadministration of antacid may affect the oral absorption and gastric adhesion of aceglutamide aluminum complex(AGA). After the oral administration of $\^$14/C-AGA and antacid to rats, the radioactivity of plasma and urinary recovery was lower than that of $\^$14/C-AGA administered group. Relatively, the cumulative recovery of radioactivity in feces was increased significantly. The comparative bioavailability of $\^$14/C-AGA from the plasma concentration-time curve and urinary recovery was about 60%. in vitro, the effect of antacid in the gastric adhesion of AGA was not significantly different between AGA and AGA/antacid treatment. And it accorded well with the result of in vivo experiment. In conclusion, on the combination of antacid, the oral absorption of AGA was decreased but the gastric adhesion was not affected in respect of drug interaction.

독사 또는 곤충의 독 30여종을 대상으로 SNU-1 위암세포에 대하여 MTT test를 실시한 결과 세포 독성 활성이 제일 높은 킹코브라(Ophiophagus hannah)의 venom을 가지고 세포 독성 물질을 정제하였다. Gel Filtration Chromatography와 Anion Exchange Chromatography로 정제한 4번째 peak만이 MTT/SRB test결과 IC$_{50}$ value 0.947$\mu\textrm{g}$/ml이었다. 음이온 교환 크로마토그라피로 정제한 단백질을 PRO-RPC로 더 분리하여 순수한 단일성분을 얻었으며 맹장암, 대장암, melanoma, fibrosarcoma 세포에 대해 독성을 확인하였고, 광학 및 전자 현미경에 의해 암세포의 분화와 성장이 억제됨을 재확인하였다. 또한 thymidine uptake asaay에서 암세포의 증식이 억제되었고, 또한 EDTA, $Zn^{++}$, $Ba^{++}$ 첨가로 세포 독성 활성이 증가되었다. (중략)

In vivo 실험결과, FVA가 감염된 BALB/c mice 에게 AZT를 18일간 경구투여시 (100mg/kg/day) 대조군에 비해 비장의 비대를 82% 억제하였으며, 이들에게 FVA를 재감염시킨 후 18일 뒤에도 비장의 비대가 63% 억제되었다. 그러나 혈액중 빈혈의 지표는 정상으로 회복하지 못하였다. 한편, tannic acid 는 (500mg/kg/day) 비장 비대를 55% 억제하였으며, 빈혈지표도 정상으로 회복되었다. 그러나, FVA를 재감염시킨 경우에는 비장의 비대가 억제되지 않았다. 반면에, tannic acid 와 AZT 를 병용투여시 비장의 비대를 98% 억제하여 상승효과를 초래하였으며, 빈혈지표도 정상으로 회복되어 독성을 감소시켰다. 또한 이들에게 FVA를 재감염시킨 경우에도 비장의 비대가 63% 억제되었다. 한편, In vitro 실험결과 AZT 의 경우 $IC_{50}$/이 0,01$\mu$M인 반면에 tannic acid의 경우에는 5 $\mu$M로서 약 1/500 의 효력을 나타내었다. 이상의 결과는 tannic acid 는 AZT 와는 다른 기전에 의해 FVA 의 감염에 의한 비장비대 및 빈혈을 억제하는 것으로 고려된다.

Several behavioral studies have suggested that ginseng total saponin (GTS) antagonizes morphine actions. Based on these observations, we conducted biochemical studies to elucidate the cellular mechanism of GTS actions. morphine hydrochloride (10mg/kg, sc) and/or on (400mg/kg, oral ) were administered to mice for 14 consecutive days. Ligand binding studies were conducted from striatal membranes. For opioid receptors, morphine increased the affinity but decreased the maximal binding sites for $^3$H naloxone. GTS partially recovered it. In case of dopamine receptors, morphine increased affinity and maximal binding sites for 3H spiperone. and GTS partially blocked it. These results suggest that morphine affects cellular events by modulating opioid receptors and that opioid receptors interact with dopamine receptors to change the mental status. GTS could be helpful for the treatment of morphine- induced mental disorders.

The effects of ginseng prepation, Adaptagen$\^$R/ (AD), on the central dopaminergic nervous system in the learning-impaired rats were studied. The learning impaired rats were rendered by the intracerebroventricular infusion of ethylcholine aziridium (AF64A), 3 nmol/each side. Three days after the infusion of AF64A, AD were orally intubated daily for five days, 200 mg/kg. The control groups were intubated with distilled water. Twenty four hours after the last intubation, The changes in the specific bindings of dopamine receptors, the concentrations of dopamine (DA) and metabolites, The activities of tyrosine hydrosylase (TH) and monoamine oxidase (MAO) were analyzed using receptor radiography, HPLC-ECD and the methods in enzyme-assays, respectively.

The present study was undertaken to investigate the behavioral and biochemical effects of ginseng total sponin (GTS) on methamphetamine-treated mice. GTS (50 or 100 mg/kg) was administered intraperitoneally two times with 2 hour interval. Two hours after the second injection of GTS, methamphetamine (2 mg/kg) was administered subcutaneously. The ambulatory activity of mice was measured by the ti1ting-type ambulometer every 10 min. for 1 hour. Methamphetamine-induced hyperactivity was reduced by GTS. in a dose-dependent manner. To study the neurochemical mechanism underlying the GTS effects, monoamine contents were measured from brain tissues. After 45 min. of methamphetamine injection. mice were sacrificed and monoamine contents were determined from the striatum. Biochemical analysis revealed that GTS reduced the methamphetamine- induced increase in striatal dopamine contents. These observations indicate that inhibition of methamphetamine-induced hyperactivity by GTS is mediated by the modulation of dopaminergic nervous system, and it could be helpful for the therapy of hyperactivity.

There have been several reports regarding the effects of Panax ginseng on allergy reactions. However, they are very sporadic and no systemic yet. To study the effects of Panax ginseng on hypersensitivity, either ginseng total saponin (GTS, 200mg/kg, oral, two hours prior to experiments) or ethanol extract (50 and 200 mg/kg, oral, one week) was administered. Various parameters were employed to assess the anti-allergic actions of Panax ginseng 48hr passive cutaneous anaphylaxis (PCA), skin reactions, histamine release from rat peritoneal mast eel Is, and lipoxygenase activity. In 48hr PCA, and in skin reactions induced by chemical mediators (histamine, serotonin) and mediator releaser (compound 48/80), Panax ginseng did not suppress sensitized immune functions, rather showed tendency to increase the histamine-induced vascular permeabi1ity. Panax ginseng did not inhibit lipoxygenase activity either.

Morphine produces wide-spread immunesuppression, such as lymphokine production, phagocytic activity, natural killer cell activity, cell proliferation, and cell-mediated immunity. On the other hand, one of the representative pharmacological act ions of Panax ginseng is immune enhancement. In this study, we investigated the effects of ginseng total sponin (GTS), and Adaptagen (trade name of ginseng product) on morphine-induced immunesuppression. Morphine hydrochloride (10 mg/kg, sc), GTS (400 mg/kg, oral), Adaptagen (400 mg/kg, oral ) were administered to mice for 14 consecutive days.

새로운 간장질환 치료제로 개발중인 YH439가 orotic acid, nicotinamide 및 ethionine에 의해 유발되는 지방간에 대한보호 및 치료효과를 관찰하였다. 웅성 SD계 rat에 orotic acid(1%) 또는 nicotinamide(2%)가 첨가된 사료를2주간 섭취시켜 유발된 지방간에 대한 YH439의 보호 및 치료효과와 ethionine(100mg/kg, i.p.)투여로 유도되는 지방간에 대한 YM439의 보호효과를 관찰하기 위하여 간 조직 중 triglyceride, cholesterol 및 phospholipid의 함량을 측정하여 비교하였다. 그 결과, erotic acid 또는 nicotinamide의 섭취로 인해 랫드의 간 조직 중 triglyceride, cholesterol 및 phospholipid의 함량은 1.5-3배정도 증가하였으며, 이 증가된 지질들은 YH439 100, 200mg/kg 투여에 의해 유의성 있게 억제되었다. 또한 ethionine투여에 의해서도 랫드의 간 조직 중 triglyceride, cholesterol 및 phospholipid의 함량은 약 2배정도 증가되었으며, YH439 전저치에 의해 이 현상들이 억제되는 효과를 나타내었다.

A polysaccharide, G009, isolated from Ganoderma lucidum IY009 subjected to investigating on general pharmacology. This material at the large oral doses of 1000 and 2000mg/kg in mice did neither exhibit any abnormal behaviors nor effects on central nervous system. It also had no influences on hexobarbital-induced sleeping time, rotarod test and spontaneous activity test at each oral dose of 1000mg/kg in mice. No effects on the body temperature and on acetic acid induced writhing syndrome in mice were observed with its oral administration at 1000mg/kg, and the convulsions induced by strychnine and pentetrazole were not inhibited at its oral doses of 1000mg/kg in mice. The solution of G009 as given intravenously at the doses of 30 and 60mg/kg in rabbit had no influences on blood pressure and respiration rates and depth. In isolated organs of rat uterus and fundus muscles and guinea-pig ileum and trachea, it did not show any contraction or relaxation at the concentration of 2$\times$10$^{-3}$g/ml, and the contractive actions produced by oxytocin, acetylcholine, serotonin and histamine did not inhibited by the same doses. This material showed no effect on intestinal propulsion test in mice and gastric secretion in rats at the oral doses of 1000mg/kg. However, it is interesting that the material exhibited potent inhibition of acidified aspirin induced gastric damage at the doses of 500 and 1000mg/kg in rats.

The purpose of this study was to observe the effects of the polysaccharide(G009) obtained from liquid cultured Ganoderma lucidum IY009 on the lipidperoxidation in rat liver microsome. It is well known that the polysaccharide of G. lucidum have the hepatoprotective activity, antitumor activity etc., which was thought to have the relationship to anti-lipidperoxidation. In order to the estimate the effects of anti-lipidperoxidation of the polysaccharide obtained from G. lucidum IY009, enzymatic and nonenzymatic reaction were performed, in vitro, in rat liver microsome. In enzymatic lipid peroxidation reaction by ADP/FeCl$_3$/NADPH and $CCl_4$/NADPH, G009(1mg/ml) inhibited 77.4%, 39.4%, respectively, and the nonenzymatic reaction strongly exhibited 97.4% inhibition. And also, in enzymatic and nonenzymatic inducers treated with G009, the formation of MDA was progressively greater decreased by raising G009 concentration. These results suggest that anti-lipidperoxidation by G009 treatment may be play an important part in liver protection action.

Pharmacokinetic studies on time-course of blood levels, tissue distribution, and excretion of G009, a potential hepatoprotective agent, were performed in male rats after a single oral dose(20mg/kg) of $\^$14/C-labelled G009. The radioactivity concentrations in plasma during 0～3 hours are low, but subsequently increase to a maximum at 12 hours after dosing. $\^$14/C-G009 was well distributed to all tissue. Tissue concentration profiles of radioactivity vary among tissues on time-course after administration. G009(single oral dosage) was distributed and/or absorbed at gastric intestines and excretional organs for initial time of 0-7 hours, and distributed to most tissue at 12-24 hours. In special, the concentration of radioactivity in tiller at 48 hours were 1% of total radioactivity of $\^$14/C-G009 administered. The expired air, urinary and fecal excretion of radioactivity within 24hours after administration were 61.5%, 1.9% and 21.2% of total radioactivity of $\^$14/C-G009 administered. The biliary excretion of radioactivity in rat increased slightly for 0-6 hours after administration. The biliary excretion of radioactivity within 48hours were 1.97%.

정상 동물군의 GOT값 및 GPT 값이 각각 137.8$\pm$23.4 및 67.4$\pm$10.0 이었고 acetaminophen 투여군은 418.6$\pm$69.1 및 447.4$\pm$7.7로 증가하였으나 G009 투여군은 그 증가가 현저히 억제되었다. 즉 G009 10mg/kg 투여군은 GOT 및 GPT 값이 192.6$\pm$19.2 및 144.8$\pm$28.7에 그쳐 각각 GOT 및 GPT 값의 상승이 80.5% 및 79.6% 억제되었고 20mg/kg 투여군은 90.8% 및 86.6% 억제되었다. 그러나 Licovek 은 50mg/kg의 투여군에서 이와 유사한 효과가 관찰되었다. 또한 간 절편의 조직학적 관찰 결과 acetaminophen 투여 대조군이 간세포 괴사등 심한 조직 괴사를 보인 반면 G009 10mg/kg 및 20mg/kg 농도에서는 간장 손상이 매우 경미함을 확인할 수 있었다.

This study was carried out to investigate the dose dependent antifibrotic effects of G009, the water-soluble fraction of polysaccharide extracted from Ganoderma lucidum. The experimental hepatic cirrhosis was induced by bile duct ligation/scission (BDL/S) in rats. BDL/S rats in each group were dosed 0.5, 2, 5 or 10mg/rat/day orally for 4 weeks after the operation. Antifibrotic effects were evaluated by serum biochemical values, hydroxyproline contents, and light microscopical histology.

1. 혈압 및 맥박 실험 Acetylcholine과 상엽 수층분획을 단독으로 정맥에 투여한 경우 각각에서 농도 의존적으로 일시적인 혈압 강하 효과가 나타났고 빈맥 현상이 관찰되었다. Nitric Oxide 합성효소 억제제인 N$^{G}$ -Nitro-L-Arginine Methyl Ester(10 mg/kg I. v)를 전처리한 경우 위 두 약물에 의한 혈압강하효과는 공히 증가되어졌다. 또한 두 약물에 의한 혈압강하 효과는 Atropine Sulfate(1 mg/kg i.v) 전처리로 완전히 차단되었다. Cholinesterase 억제제인 Physostigmine (0.05 mg/kg i.v) 전처리는 상엽의 혈압강하 효과에 아무런 영향을 나타내지 못하였다. 2. 장관 실험 Acetylcholine과 상엽 수층분획을 organ bath에 첨가한 경우 각각에서 농도 의존적으로 장관 수축력을 증가시켰다. 혈압반응에서와 같이 장관실험에서도 두 약물에 의한 장관 수축력의 증가는 Atropine Sulfate(1$\times$$10^{-5}$ M)의 존재하에서는 완전히 차단되어졌다. 이상의 결과로부터 상엽 수층분획은 Acetylcholine과 유사한 작용을 지닌 물질을 함유하는 것으로 사료된다.

간장 손상시에는 여러 혈청의 효소 활성과 함께 혈청 $\beta$-glucuronidase의 활성도 증가한다는 것이 보고되었으나 심한 간부전이나 간암의 경우 이들의 활성은 오히려 정상치보다 감소하는 것으로 나타났다. Silymarin은 간장 보호효과로 이미 임상에 널리 사용되고 있는 약물로서 김 등에 의해 silymarin이 장내세균의 $\beta$-glucuronidase와 간장의 $\beta$-glucuronidase의 활성을 억제한다는 것이 보고되었다. 이에 연자 둥은 $\beta$-glucuronidase의 저해 효과가 관찰된 영지버섯온 유기용매로 분획하여 간장 보호효과를 검색하였다. 영지버섯의 70% MeOH 추출물(GT)과 그 ether 분획(GE)에 대해 생쥐 1군을 6마리로 하여 20% $CCl_4$0.1$m\ell$/10g(olive oil로 희석)을 경구투여 하였다. 검액 GE는 50mg/kg과 250mg/kg, GT는 100mg/kg과 500mg/kg을 각각 사염화탄소 투여 30분 전에 경구투여 하였으며 사염화탄소를 투여하고 24시간 후에 심장 채혈하고 혈청을 분리하여 혈청성분 및 혈청효소의 활성을 측정하였다. 대조군에는 생리식염수를 투여하였고 양성 비교약물로는 silymarln 100mg/kg을 경구투여 하여 비교 관찰하였다. 실험 결과, 영지버섯의 ether 분획에서는 혈청중 GOT, GPT의 활성과 triglyceride의 함량에 대해 silymarin보다 우수한 효과를 보였으며, 70% MeOH 추출물은 silymarin에 미치지 못했다.

본 연구는 표고버섯에서 분리정제한 렉틴 성분(LEL)을 말초혈액 단핵세포(PBMC)에 반응시켜 사이토카인 유도능을 지질다당류(LPS)와 비교하여 역전사효소 중합반응법(RT-PCR)으로 측정하였다. 측정 대상 사이토카인은 IL-1, IL-2, IL-6, TNF$\alpha$ 및 IFN${\gamma}$의 다섯가지였으며 이들을 대상으로 PBMC에 렉틴을 적용하여 1, 8, 24, 48, 72, 96, 120분 등의 시간대에서 반응시켜 사이토카인의 유전자 발현 유도에 관한 다음의 결과를 얻었다. LEL의 사용 농도에 따른 편도선 림프구(tonsillar lymphocyte)의 TNF$\alpha$ 유전자 발현 양상은 반응 1시간의 경우 LEL의 일부 농도와 LPS 전농도에서 관찰되었으나 반응 40시간째에는 LEL 전농도와 LPS 전농도에서 TNF$\alpha$ 유전자 발현 양상을 관찰할 수 없었다. RT-PCR 결과 원액이나 회석액 재료로부터 관찰된 TNF$\alpha$유전자 band의 강 약 차이는 나타나지 않았다. LEL의 자극에 의한 반응 시간대 별 PBMC에 의한 사이토카인 유전자 발현 양상은 위에서 언급된 다섯가지 사이토카인을 유도, 생성할 수 있다는 것이 확인되었는데, IL-2, IL-6 및 IFN${\gamma}$는 120시간까지 장시간 지속되는 유전자 발현이 가능한 반면 TNF $\alpha$의 생성 양상은 이들 사이토카인의 생성 양상과는 판이하게 반응 1, 8 및 24 시간대까지만 TNF$\alpha$ 유전자 발현을 관찰할 수 있었고 IL-1은 72 시간까지 반응을 나타내는 등 특이적 양상을 보였다. 한편 LPS는 실험에 사용된 전 사이토카인의 유전자 발현을 120시간대까지 반응이 유지됨을 관찰하였기에 LPS가 PBMC의 강력한 사이토카인 유도체임을 입증할 수 있었으며 LEL과 다소 상이한 결과를 보였으나 LEL 또한 PBMC로부터 사이토카인을 생성 유지시킬 수 있는 유도체로 작용함을 확인할 수 있었다.

이 연구에서는 혈전증 치료에 사용되는 혈전 응해제를 국내 독사독으로부터 개발하기 위한 실험을 실시하였다. 그 내용을 요약하면 다음과 같다. 국내에 서식하는 독사인 Agkistrodon blomoffi brevicaudus, Agkistrodon caliginosus와 Agkistrodon saxatilis에서 각각 사독을 채취하여 fibrin plate 방법으로 fibrin 분해능을 조사하여 Agkistrodon blomoffi brevicaudus의 독이 분해능이 가장 우수함을 밝혔다. 이와 같은 사실에 기초하여 A. blomoffi brevicaudus의 독으로부터 p-Aminobenzamidine affinity chromatography와 DEAE ion-exchange chromatography를 이용하여 분자량이 50.8 kDa인 황성 단백질을 정제하였다. 위에서와 같은 방법으로 정제한 단백질은 fibrin 분해능이 우수하고 fibrinogen의 ${\gamma}$ chain은 분해하지 않으나 B$\beta$ chain을 $A\alpha$ chain에 비하여 보다 선택적으로 분해하는 단백분해 효소임을 증명하였다. 이 정제 효소의 Fibrin에 대한 분해능은 266$\mu\textrm{g}$/${\mu}\ell$의 농도에서 Plasmin 1.0 unit (=3.0 WHO unit)보다 높게 나타났다. 정제된 효소는 chromogenic substrate인 N-Benzoyl-Phe-Val-Arg-pNA와 N-p-Tosyl- Gly-Pro-Arg-pNA의 arginine carboxyl side를 분해하고 pH 7.5에서 최대 활성을 보이며, Vmax는 5.46 umo1/1ㆍmin이고, Km 값은 0.20mM이며, 그리고 Cu$^{2+}$, $Zn^{2+}$, soybean trypsin inhibtor에 의해 25～50% 정도, serine proteinase inhibitor인 phenylmethylsulfonyl floride에 의해 80%정도 활성이 억제되는 특성이 있음을 규명하였다.

저자 등은 일련의 식물엑스에 대하여 항위염 및 항궤양효능에 관한 검색을 실시하여 오리나무수피의 MeOH엑스가 현저한 효과가 있음을 예지 하였으므로 그에 대하여 보다 구체적인 실험을 실시하였다. 즉 MeOH엑스를 Hexane, CMCl$_3$, BuOH로 계통적으로 추출하여 상기의 분획 및 잔사인 물분획을 제조하여 이에 대한 실험을 실시하였다. 소화성궤양이 공격인자와 방어인자의 불균형에 의해 형성됨을 Shay가 주장함으로써 공격인자의 억제를 알아보기 위한 Mizui 등의 방법인 HClㆍEtOH 유발 위손상 실험과 Guth 등의 Aspirin 위손상 실험 방법에 따라서 위 손상정표를 관찰하였다. Shay의 방법에 따라서 위궤양 모텔은 유문을 결찰하고 검체를 십이지장내에 투여하고 처치를 완료하고 12시간 후에 궤양정도와 또한 4시간의 유문결찰에 의한 위액분 비량, pH 및 산분비량에 미치는 영향을 관찰하였다. 또 염산과 pepsin 등에 의한 점막 손상에 대한 방어인자의 증강요인인 위 mucous membrane의 mucus분비를 알아보기 위한 absoluteㆍ에탄올 위손상에 대한 예방효과 시험을 시행하였다. 이 실험의 결과, 오리나무 MeOH엑스의 항위염 및 항위궤양효과는 BuOH 분획에서 강력한 작용이 있었으며 이 분획은 aspirin 유발 위손상 및 shay 궤양에 효과를 나타내었고 또한 mucin량의 증가를 보여주었다. 그러나 이 분획은 위액, pH 및 산분비량에 영향을 나타내지 아니하였다.

위염(gastritis), 위궤양(gastric ulcer)은 소화기 질환중에서 가장 빈도수가 높은 질병으로 위장관 점막이 위산에 의해 소화되어 버리므로서 궤양을 형성하는 상태를 말한다. 위염 및 위궤양의 발생빈도가 높은 것에 비해 발생원인은 정확히 밝혀져 있지 않으며 대체로 공격인자와 방어인자의 불균형 즉, 공격인자의 증가나 방어인자의 약화 또는 세균에 의한 감염에 의해 발생되는 것으로 알려져 있다. 일련의 식물엑스에 대하여 항위얌 및 항궤양효능에 관한 검색을 실시하여 노두의 M？H엑스가 현저한 효과가 있음을 예지하였으므로 그에 대하여 보다 구체적인 실험을 실시하였다. 즉 MeOH 엑스를 Hexane, CHCl$_3$, BuOH로 계통적으로 추출하여 상기의 분획 및 잔사인 물분획을 제조하여 이에 대한 실험을 실시하였다.

This study was performed to evaluate the general toxicity of novel Pt(II) complexes, (KHPC-002: [Pt(trans-1-dach) (DPPE)].2NO$_3$, KHPC-005: [Pt(cis-dach)(DPPE)].2NO$_3$ and KHPC-006: [Pt(cis-dach)(DPPP)]. 2NO$_3$). In the acute toxicity study in rats, three dosing groups of Sprague-Dawley male rats in each compounds were given a single intraperitoneal injection of KHPC-002, KHPC-005 and KHPC-006. In order to compare the toxic effects of these novel Pt(II) complexes with those of cisplatin, one group Sprague-Dawley male rats were given 7mg/kg i.p injection of cisplatin. Body weights showed dose-related decrease in all treatment groups when compared wi th the control group.

This study was conducted to examine novel Pt(II) complexes, (KHPC-002; [Pt(trans-1-dach) (DPPE)]. 2NO$_3$, KHPC-005;[Pt(cis-1-dach)(DPPP)]. 2NO$_3$ and KHPC-006; [Pt(cis-1-dach) (DPPE)]. 2NO$_3$) for their acute toxicities and toxicological profiles in preclinical studies. In male and female mice given a single intraperitoneal administration of KHPC-002, KHPC-005 and LHPC-006, we determined that LD$\_$50/ values of pt(II) complexes were 295.5mg/kg(M), 350.4mg/kg(F);KHPC-002, 158.7mg/kg(M), 157.7mg/kg(F); KHPC-005, 574.8mg/kg(M), 596.5 mg/kg (F); KHPC-006, respectively. In gross and histopathological examination on dead animals, no abnormal changes were observed in any organs.

This study was conducted to examine novel Pt(II) complexes, (KHPC-002: [Pt(trans-1-dach) (DPPE)]. 2N0$_3$, KHPC-005: [Pt(cis-dach) (DPPE)] 2NO$_3$ and KHPC-006: [Pt(cis-dach) (DPPP)]. 2NO$_3$) for their acute toxicities. In male and female mice given a single intraperitoneal administration of KHPC-002, KHPC-005 and KHPC-006, we determined that LD$\_$50/ values of Pt(II) complexes were 295.5mg/kg(M), 350.4mg/kg(F) : KHPC-002, 596.5mg/kg(M), 674,8mg/kg(F): KHPC-005, 158.7mg/kg(M), 157.7mg/kg(F); KHPC-006, respectively. The signs of toxicity in mice observed fellowing the administration of these compounds included the followings: decreased mortor activity: abnormal gait: salviation, trasient decreased body weight. There were no treatment related specific changes in growth examination.

본 연구는 유전공학적인 방법으로 합성된 상피세포성장호르몬인 DWP-401에 대한 마우스의 반복투여에 의한 아급성독성을 조사하기 위하여 실시하였다. 시험군은 ICR 마우스 압수 각각 10 마리씩으로 하여 3 개의 처치군 및 대조군(0, 1, 0.2, 0.04 mg/kg)과 회복시험군(0, 1 mg/kg)을 두었다. 시험물질은 13 주간 경배 부 피하에 1 주에 6 회의 빈도로 투여하였고 대조군과 최고용량 군에서 5 주간의 회복기간을 두었다. 시험기간 중 체중, 사료섭취량 및 음수섭취량을 측정하였고, 뇨검사, 안검사, 혈액학적검사, 혈액생화학적 검사, 부검소견관찰, 장기중량측정 및 병리조직학적검사를 실시하였다. 이상의 실험결과 DWP-401의 마우스에 대한 표적장기는 상피세포, 간장, 비장, 부신, 방광 신장, 각 장기의 복막과 흉막, 림프계, 난소 및 투여부위였고 회복성이 인정되었다. 무해용량은 0.04 mg/kg/day였으며 확실 중독량은 0.2 mg/kg/day 이상으로 사료되었다.

To evaluate the genetic toxicity of NP-77A which is selected as the candidate of anti-HBV agent, we performed ames test, micronucleus test, and chromosome aberration test on the CHL cell in vitro. The Ames test was carried out with 5 fold diluted 5 concentrations from 25mg/plate using S. typhimurium and E.coli. After 48hrs incubation, revertant colony numbers was calculated with and without metabolic activation system. In vivo micronucleus test, we investigated the rate of the occurrence of micronucleus after I.P. administration to mice. Andalso, we observed the incidence rate of cells with chromosomal aberration by NP-77A treatment using CHL cell line.

Cisplatinum is often effective in cancer treatment, but potent nephrotoxicity limits its clinical use. We have, therefore, developed new anticancer drugs that contain platinum. We have synthesized six new platinum compounds based on Figure 1. Drugs were initially administrated at 5${\times}$10$\^$-4/M with 48 hours exposure in monolayer cultures of primary rabbit proximal tubular cells and human renal cortical cells with the M.T.T. endpoint to measure toxicity. Drug concentrations of 10$\^$-3/M, 10$\^$-4/M, and 10$\^$-5/M with 72 hours exposure were used for human renal cortical tissues in 7 weeks histoculture with toxicity measured by the glucose-consumption endpoint. From these studies, we determined that the new platinum drugs have lower nephrotoxicity than cisplatinum. Drugs D, E, and H. have lower nephrotoxicity than the other new drugs. We are currently measuring the anticancer efficacy of drugs D, E, and H.

Quinolone계 항균제인 rufloxacin의 전임상시험의 일환으로 항원성 유발여부를 평가하기 여부를 평가하기 위하여 아나필락시스 쇼크 반응시험, 수동 피부 아나필락시스 반응시험 및 간접 적혈구 응집 반응시험을 실시하여 다음과 같은 결과를 얻었다. 1. 기니픽을 이용한 아나필락시스 쇼크 반응시험에서 rufloxacin의 저용량투여군(1mg/body, 추정임상용량), 고용량투여군(5mg/body), 고용량혼합투여군(5mg/body mixed with Freunds' complete or incomplete adjuvant) 및 단백결합혼합투여군(rufloxacin-BSA, 1mg/body)에 있어서 아나필락시스 쇼크 반응이 관찰되지 않았다. 2. 마우스-랫드를 이용한 수동 피부 아나필락시스 반응시험에서 저용량투여군, 고용량투여군, 오용량혼합투여군 및 bovine serum albumin(BSA)과 결합시킨 단백결합혼합투여군 모두에서 음성의 반응을 나타내었다. 3. 간접 적혈구 응집 반응시험 결과 저용량투여군, 고용량투여군, 고용량혼합투여군 및 단백결합혼합투여군에서는 각군당 1-4마리에서 16-32배에서 양성반응을 나타내었다. 그러나 음성대조군에서도 10마리 중 4마리에서 16-32배에서 양성반응을 나타낸 것으로 미루어 약물투여군에서의 미약란 양성반응이 약물특이적 반응은 아닌 것으로 사료된다. 이상의 실험결과로 미루어 rufloxacin은 항체 생성능은 가지지 않는 것으로 사료된다. 이상의 실험결과를 종합하여 볼 때 ndloxacin은 전반적인 항원성 potential은 없는 것으로 사료되며 또한 생체내에서 체내 단백과 결합하여 hapten으로 작용할 가능성은 없는 것으로 사료된다.

본 연구는 중금속에 의한 생체내 독성기전을 연구하고자 흰쥐간의 상층액에서 카드뮴과 잘 결합하는 HMWP들을 분리, 정재하고 나아가 생화학적 특성을 밝히고 이단백질이 카드뮴에 의해 생성되는 것인지 아니면 간의 기존 단백질인지를 밝히고자 함을 연구의 목적으로 하였다. CdCl %2 (3mg/kg body wt.)을 3일간 ip injection시킨 후 흰쥐의 간을 적출하고 균질화하여 원심분리한 crude extract를 직접 Sephacryl S-100에 흡착시켜 10mM phosphate buffer(pH 7.0)으로 용출시켰다. 용출된 fraction tube를 UV spectrometer로 흡광도를 측정하고 atomic absorption spectrophotometer로 카드뮴량을 측정하여 카드뮴량이 높은 분획을 모아서 ion exchange column chromatography(DEAE-Sepharose)에 흡착시켜 염농도 구배로 chromatography를 실시하고, 음이온 교환수지에서 흡착되지 않고 용출된 분획을 ultrafiltration으로 농축시킨 후 S-Sepharose에 흡착시켜 염농도 구배로 chromatography를 실시한 결과 두 종류의 카드뮴 결합 단백질(Cd-BP)를 분리, 정제하였다.

The present study was conducted to examine the effect of recombinant bovine somatotropin(${\gamma}$ bST), which was administered to cow to promote milk production, on bST levels in milk. Fourteen cows were divided into two groups: 1) control cows received neither y bST nor vehicle, 2) treated cows were administered twice at two-week interval with 500mg ${\gamma}$ bST each cow by subcutaneous injection. Milk samples were taken on day 0 (prior to injection), day 7 (7 days after 1st injection), day 21 (7 days after 2nd injection) and day 35 (21 days after 2nd injection).

To investigate the mechanism of the regulation of cytochrome P450IA1 gene expression, ethoxyresorufin deethylase(EROD) and benzo(a)pyrene hydroxylase in B6 mouse liver, in isolated perfused rat liver system. and in B6 mouse hepatocyte Hepa-I cells were examined. In C57BL/6N mouse, 3-methylcholan- throne( 3MC ) treatment have resulted in the stimulation of EROD activity based on fluorometry by 2.79 fold comparirng with that of control. Measurement of mRNA of cytochrome P450 was carried out by either nothern blot or dot blot analysis. Findings are similar to that of studies with enzymes. Furhtermore, when RTPCR method was applied to detect mRNA in Hepa I cell and liver tissues the results were more clear. Cytochrome P450IA1 upstream DNA containing CAT construct was transfected into Hepa-1 cells. After transfection of CAT construct, 3MC and flavonoids, such as, chrysin, hesperetin, kaempferol, morin, myricetin and aminoyrine were treated. 48 Hours after treatments, cells were harvested and assayed for CAT mRNA by RTPCR. 3MC treatment to hepa I cells transfected with trout P450IA1-CAT construct increased CAT mRNA by 2.81 fold when it was compared with that of control. This increase CAT mRNA was decreased by concomitantly treated flavonoids and aminopyrine. The level of CAT protein was 29.2-58.0% of 3MC stimulated CAT protein. Results of this study suggested that RTPCR seems to be a very good method to study regulation of gene expression in liver tissue or Hepa cells.

ASPS against TGF-${\beta}$ is developing as scar formation inhibitor. The scar was caused by undesired collagen deposition due to overexpression of TGF-${\beta}$ in wounded tissue. The in vitro percutaneous absorption of ASPS(25mer)was investigated by using Furanz Diffusion Cell. The flux of ASPS cannot be found through normal skin due to high molecular weight (MW 10,000) and polyanionic charge. However, the skin permeation of ASPS through tape-stripped damaged skin was markedly increased. The skin fluxs of ASPS were decreased in the following order; hairless mouse> rat >human cadaver skin.

AM, SM 및 ASA는 수용액중에서 겉보기 1차반응에 따라 분해되었으며 보존온도가 높을수록 분해가 촉진되는 온도 의존성을 나타내었다. AM의 분해경로는 pH 1.22 및 pH 7.0 이상에서는 AM$\longrightarrow$ SM $\longrightarrow$ SA의 경로로 주로 분해되었으며 pH 2.01 - 6.08의 범위에서는 AM $\longrightarrow$ASA$\longrightarrow$SA의 경로로 분해되는 양상을 보였다. 또 pH가 분해에 미치는 영향을 pH-rate profile로 나타낸 결과 AM, SM 및 ASA의 최대안정 pH는 각각 4.0, 3.0, 2.0 부근이 있고 이 조건에서의 분해 반감기는 114, 168, 113 hr로 나타났다. 전체적으로 보면 pH 2.0 이하에서는 ASA가 AM 보다 약간 안정한 편이나 pH 2.0-8.0 사이에서는 AM의 분해속도가 ASA보다 현저히 낮았다. 또 AM은 pH 7.0 이상에서, SM은 pH 6.0 이상에서, ASA는 9.0 이상에서 특수염기촉매반응에 따라 분해가 이루어지는 것을 알 수 있었다. 이온강도($\mu$)의 영향으로는 pH 7.0에서 이온강도가 0.115에서 1.0으로 증가할수록 $\mu$$^{1}$2/에 대해 AM의 분해속도정수가 직선적으로 완만하게 감소되었다. 또 완충수용액 중 AM의 가수분해 억제효과를 검토하기 위해 시클로덱스트린류를 첨가하였을 때, $\beta$-시클로덱스트린과 히드록시프로필기-$\beta$-시클로덱스트린은 AM의 분해를 각각 1.6배 및 1.1배 촉진시켜 촉매적으로 작용하였으며 디메칠-$\beta$-시클로덱스트린은 약 3.2배 분해속도를 억제시켜 안정화제로 작용하였다.Zn^{2+}$, soybean trypsin inhibtor에 의해 25～50% 정도, serine proteinase inhibitor인 phenylmethylsulfonyl floride에 의해 80%정도 활성이 억제되는 특성이 있음을 규명하였다.면역환성 (immunoreactivity)이 나타났고 pyramidal cell layer (PCL)와 glia에 SOD-1이 강하게 염색되었다. APT 병용 투여로 상당수의 경련이 일어나지 않은 흰쥐는 해마의 DG에 FRA가 경미하게 염색되었고, PCL에 SOD-1도 경미하게 나타났으나, 경련이 나타난 쥐에서는 KA만을 투여한 흰쥐와 구별되지 않았다. 이상의 APT의 항산화 효과는 KA로 인한 뇌세포 변성 개선에 중요한 인자로 작용할 것으로 사료되나, 보다 명확한 APT의 기전을 검색하고 직접 임상에 응응하기 위하여는 보다 다양한 실험 조건이 보완되어야 찰 것으로 생각된다. 항우울약들의 항혈소판작용은 PKC-기질인 41-43 kD와 20 kD의 인산화를 억제함에 기인되는 것으로 사료된다.다. 것으로 사료된다.다.바와 같이 MCl에서 작은 Dv 값을 갖는데, 이것은 CdCl$_{4}$$^{2-}$ 착이온을 형성하거나 ZnCl$_{4}$$^{2-}$ , ZnCl$_{3}$$^{-}$같은 이온과 MgCl$^{+}$, MgCl$_{2}$같은 이온종을 형성하기 때문인것 같다. 한편 어떠한 용리액에서던지 NH$_{4}$$^{+}$의 경우 Dv값이 제일 작았다. 바. 본 연구의 목적중의 하나인 인체유해 중금속이온인 Hg(II), Cd(II)등이 NaCl같은 염화물이 함유된 시료용액에 공해이온으로 존재할 경우 흡착에 의한 제거가 가능하다. 한편 이같

제제설계 및 컴퓨터 최적화기법을 응용하여 고형으로 바뀌는 Gel화 시간이 가장 짧은 기본처방을 구하고, Indomethacin을 주 약물로하여 약물방출에 영향을 줄 수 있는 PVA, PEG 및 Ethanol 을 독립변수로, 약물방출속도를 종속변수로 하여 중심 합성계획법에 따라 실험을 행하여 최적처방을 구한다. 최적처방에 의한 Soft Hydrogel 을 제조하여 약물방출속도 및 Rheometer 에 의한 유동특성을 측정하였다.

OMPED의 결합비는 1:1 몰비로 복합체가 형성됨을 확인하였다. 수용액중(pH 7.0)에서 OMP는 분해속도 k=1.542$\times$$10^{-2}$$hr^{-1}$, shelf life=6.81hr 이었고 OMPED는 k=2.088$\times$$10^{-4}$$hr^{-1}$, shelf life=502.8hr이며 중성은 물론 약산성에서 안정하였다. HPMCP로 장용피한 OMPED pellet은 산저항성이 완벽하고 용출속도가 양호하였으며, OMPED 정제에서는 CAP로 코팅한 정제가 가장 큰 AUC값을 나타내었다. Witepsol H-15 기제를 사용한 각 좌제의 생체이용율은 OMPED 좌제가 86.53 $\mu\textrm{g}$ㆍmin/$m\ell$ 로서 OMP 좌제 61.9 $\mu\textrm{g}$ㆍmin/$m\ell$ 및 OMP-$\beta$-CD 좌제 68.8 $\mu\textrm{g}$ㆍmin/$m\ell$이다.

각종 OMP 복합체들은 OMP에 비해 용해도가 2.7-12.0배 증가하였으며, 모두 10분 이내에 85% 이상이 용출되어 용출규정에 적합하였고 pH 및 습도에 따른 안정성 결과도 OMP에 비해 각종 OMP 복합체가 안정성이 증가되었다. OMP-cholestyramine 수지염의 경우 온도, 습도 및 수용액 중에서의 안정성 모두 OMP에 비하여 향상되었으며, 4$0^{\circ}C$, RH 75%의 가혹조건에서도 OMP pellet에 비해 OMP-cholestyramine 수지염 pellet이 안정성과 내산성이 매우 우수하였다.