Structure-activity relationships of N-(3,5-dimethoxy-4-n-octyloxycinnamoyl)-N'-(3,4-dimethylphenyl)piperazine and analogues as inhibitors of acyl-CoA: cholesterol O-acyltransferase.

A novel series of acyl-CoA: cholesterol O-acyltransferase (ACAT) inhibitors were synthesized from a lead compound, 1-(4-hydroxy-3-methoxyphenyl)-7-phenylhept-1-en-3-one (1, Yakuchinone B) through a modification of three regions (A, B, C) in the molecule. In this study, the compounds prepared were tested for in vitro inhibitory activity on microsomal ACAT from the liver of rats and for in vivo hypocholesterolemic activity in rats given a high cholesterol diet. N-(3,5-Dimethoxy-4-n-octyloxycinnamoyl)-N'-(3,4-dimethylphenyl)piperazine (45), which belongs to the amide compounds, has finally been discovered. Compound 45 inhibited rat hepatic ACAT in a more striking manner than CI-976, an amide compound ACAT inhibitor, and it exhibited a high level of hypocholesterolemic activity in vivo. Since 45 strongly inhibited both microsomal ACAT prepared from HepG2 (a cell line derived from human hepatocarcinoma) and Caco2 (a cell line derived from human colon adenocarcinoma), there is speculation that 45 might have the ability to inhibit ACAT in both the human intestine and liver independent of the difference in the distribution of ACAT isozymes. On the other hand, 45 did not induce adrenotoxicity in subacute toxicity studies in rats. These results suggest that it has promise for development as a new therapeutic agent for hypercholesterolemia and atherosclerosis.

Effect of oligofucose derivatives on acetic acid-induced gastric ulcer in rats.

This study attempted to enhance the anti-ulcer activity of fucoidan from Cladosiphon okamuranus TOKIDA by chemical modification with a hydrophobic group. The suitable number of fucose residues in the effective compound was also clarified to obtain a compound of constant quality. Degraded fucoidans were coupled with several hydrophobic groups via Schiff bases, and their anti-ulcer activities were determined by acetic acid-induced ulcer models in rats. Size-fractionated oligofucose was also modified and assayed for anti-ulcer activity. Among the modified oligofucoses, only the oligofucose-dodecylaniline combination (OFDA) significantly promoted ulcer healing. The effective dose was 0.2 mg/kg/d. The most suitable number of fucose residues in the compound for the anti-ulcer activity was determined to be around 12. We succeeded in enhancing the anti-ulcer activity of Cladosiphon fucoidan by modification with dodecylaniline. The activity of this compound was comparable or greater than that of typical anti-ulcer agents. By determination of the optimal OF chain length for the anti-ulcer activity of OFDA, it became possible to obtain OFDA of constant quality.

Diarylheptanoids suppress expression of leukocyte adhesion molecules on human vascular endothelial cells.

Diarylheptanoids possess potent anti-inflammatory properties. However, the mechanism of their action is not fully understood. In this study, we found that three diarylheptanoids, 1-(3, 5-dimethoxy-4-hydroxyphenyl)-7-phenylhept-1-en-3-one (YPE-01), yakuchinone B and demethyl-yakuchinone B, reduced the adhesion of both human monocytic cell line U937 and human eosinophilic cell line EoL-1 cells to tumor necrosis factor-alpha (TNF-alpha)-treated human umbilical vein endothelial cells. In addition, they suppressed interleukin-1beta- or TNF-alpha-induced expression of E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) on the surface of the endothelial cells. Since YPE-01 reduced both VCAM-1 and ICAM-1 mRNA induction in TNF-alpha-stimulated endothelial cells, diarylheptanoids appeared to suppress adhesion molecule expression at the transcriptional level. Furthermore, YPE-01 suppressed both VCAM-1 and ICAM-1 mRNA induction as well as edema in 12-O-tetradecanoylphorbol 13-acetate (TPA)-inflamed mice ears in vivo. These results suggest that the anti-inflammatory action of diarylheptanoids is, at least in part, due to their suppressive effect on the surface expression of inducible adhesion molecules in endothelial cells, and subsequent leukocyte adhesion.

Properties of fucoidan from Cladosiphon okamuranus tokida in gastric mucosal protection.

To elucidate the anti-ulcer potential of Cladosiphon fucoidan, anti-peptic activity, bFGF stabilizing activity and inflammatory properties of this and related substances were investigated. Anti-peptic activity was observed with this and other sulfated polysaccharides such as dextran sulfate, carrageenan, and Fucus fucoidan. However, non-sulfated polysaccharides such as mannan and dextran did not exert the anti-peptic activity. The loss of bFGF bioactivity was prevented by all sulfated polysaccharides tested except chondroitin sulfate, at pH 7.4 and at pH 4.0. At pH 2.0, only heparin protected the bFGF activity. The generation of superoxide by macrophages and PMNs was stimulated by dextran sulfate, carrageenan, and Fucus fucoidan, whereas Cladosiphon fucoidan, heparin and chondroitin did not. Dextran sulfate, carrageenan, and Fucus fucoidan also stimulated the secretion of TNFalpha from macrophages, while Cladosiphon fucoidan did not. Thus, Cladosiphon fucoidan is a sulfated polysaccharide without inflammatory action. These results suggest that Cladosiphon fucoidan is a safe substance with potential for gastric protection.

Structural study of fucoidan from Cladosiphon okamuranus TOKIDA.

A structural study was carried out on a fucoidan isolated from the brown seaweed Cladosiphon okamuranus. The polysaccharide contained fucose, glucuronic acid and sulfate in a molar ratio of about 6.1 : 1.0 : 2.9. The results of Smith degradation showed that this polysaccharide has a linear backbone of 1-->3-linked alpha-fucopyranose with a half sulfate substitution at the 4-positions, and a portion of the fucose residues was O-acetylated. The data obtained from partial acid hydrolysis, a methylation analysis and NMR spectra indicated that the alpha-glucuronic acid residue is linked to the 2-positions of the fucose residues, which were not substituted by a sulfate group. These results indicated that the average structure of this fucoidan is as follows: -[(-->3Fuc-4(+/-OSO3-)alpha1-)5-->3[GlcA alpha1-->2]Fuc alpha1-]n-. (Half of each fucose residue was sulfated. One O-acetyl ester was present in every 6 fucose residues.)

Anti-inflammatory effect of YPE-01, a novel diarylheptanoid derivative, on dermal inflammation in mice.

OBJECTIVE AND DESIGN: We investigated the anti-inflammatory effect of YPE-01, a novel diarylheptanoid derivative in vitro and in vivo. MATERIAL: In the in vitro study, rat basophilic leukemia (RBL-1) cells were used. For the in vivo study, ICR and ddY mice (male, 7 weeks old) were used. TREATMENT: In the in vitro study, the supernatant of RBL-1 cells lysate was incubated with 50 microM arachidonic acid (AA) and 0.01-100 microM test drugs for 15 min. RBL-1 cells were preincubated with 0.01-100 microM test drugs for 10 min before incubation with 0.5 microM calcium ionophore A23187 for 10 min. In the in vivo study, YPE-01 (0.1-3 mg/ear) was applied to the ear of mice at the same time as a 12-O-tetradecanoylphorbol 13-acetate (TPA) application or 1 h before an AA application. METHODS: In the in vitro study, the amounts of 5-hydroxyeicosatetraenoic acid and leukotrienes were measured by high-performance liquid chromatography and an enzyme immunoassay, respectively. In the in vivo study, a circular tissue sample from the ear of the mice was weighed. Statistical analysis was done using Dunnett's test. RESULTS: YPE-01 inhibited the 5-lipoxygenase activity (IC50, 0.28 microM) and the leukotriene B4 (IC50, 0.035 microM) and C4 (IC50, 0.046 microM) production by RBL-1 cells without any inhibition of cyclooxygenase activity in vitro. The topical application of YPE-01 significantly suppressed both the AA- and TPA-induced ear edemas in vivo. CONCLUSIONS: YPE-01 is a selective 5-lipoxygenase inhibitor with a suppressive effect against dermal inflammation.

Tyrosinase inhibitors from crude drugs.

Various crude drugs were examined for their tyrosinase inhibitory activity. Marked activity was observed in Chouji and Yakuchi extracts and the active substances in these extracts were identified as eugenol and yakuchinone A, respectively. Other vanillyl compounds such as ferulic acid, curcumin and yakuchinone B also had higher activities than eugenol or yakuchinone A and inhibited the enzyme competitively. The presence of the hydroxyl group at the 4 position of the aromatic ring of the cinnamoyl moiety and the alpha,beta-unsaturated carbonyl conjugated with an aromatic ring in these substances may play important roles in the competitive inhibition of tyrosinase.

Synthesis and antitumor activity of 20(S)-camptothecin derivatives: carbamate-linked, water-soluble derivatives of 7-ethyl-10-hydroxycamptothecin.

Novel 36 derivatives (6), bonding the phenolic hydroxyl group of 7-ethyl-10-hydroxycamptothecin (4) with diamines through a monocarbamate linkage, were synthesized and their antitumor activity was evaluated in vivo. The derivatives were soluble in water as their HCl salts with the E lactone ring intact and exhibited significant antitumor activity. One of the derivatives, 6-27 showed excellent activity against L1210 leukemia and other murine tumors. The structure of its hydrochloride trihydrate (CPT-11) was determined by spectroscopic and crystallographic methods.

Antihepatotoxic actions of gingerols and diarylheptanoids.

The antihepatotoxic effects of gingerols, shogaols, diarylheptanoids and related analogues were assessed utilizing carbon tetrachloride- and galactosamine-induced cytotoxicity in primary cultured rat hepatocytes. Most congeners exhibited significant actions in these assay methods. The carbon tetrachloride assay appeared to be most useful in defining structure-activity relationships. The antihepatotoxic activity of gingerols and shogaols was dependent on the length of the side chain with the [7]- and [8]-companions eliciting the strongest activity. The gingerols exerted more intense activity than the corresponding shogaols. In the diarylheptanoids, introduction of hydroxyl groupings on the phenyl rings caused increased activity; however, the effect of the positions and number of hydroxyls on activity was variable depending on the carbon skeleton.