Inhibition of collagen hydroxylation by lithospermic acid magnesium salt, a novel compound isolated from Salviae miltiorrhizae Radix.

We have screened several chinese medicinal herbs for the presence of antifibrotic agents. An aqueous extract of Salviae miltorrhizae Radix was found to inhibit collagen secretion by human skin fibroblasts without affecting DNA or noncollagen protein synthesis. We have subsequently purified the material exhibiting the inhibitory activity and identified it as magnesium lithospermate. From its chemical structure this compound was predicted to be an inhibitor of the post-translational modifying enzymes prolyl and lysyl hydroxylases in collagen biosynthesis. Accordingly, it decreased the extent of prolyl and lysyl hydroxylations in collagen by approx. 50%. Added to cell extracts it inhibited both prolyl and lysyl hydroxylase activities, but only lysyl hydroxylase activity when added to intact cells. Oral administration of this compound to mice led to a significant reduction of prolyl hydroxylation in newly-synthesized skin collagen. This naturally-occurring compound thus offers a potential means for treating fibrotic diseases, such as systemic scleroderma and keloid.

Study on the inhibitory effect of tannins and flavonoids against the 1,1-diphenyl-2 picrylhydrazyl radical.

Fifty-one tannins and forty-one flavonoids isolated from Oriental medicinal herbs were evaluated for their antioxidant ability with a 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-generating system. The results showed that tannins and certain flavonoids are potential free-radical scavengers, and that their activity against the DPPH radical is closely associated with their chemical structure. A comparison of the two classes of compounds showed that tannins have more potential than flavonoids because almost all the tannins demonstrated significant scavenging action within a low concentration range, whereas the activity of flavonoids varied distinctively among the different compounds. An increase of galloyl groups, molecular weight, and ortho-hydroxyl structure enhanced the activity of tannins, whereas the number and position of hydroxyl groups were important features for the scavenging of free radicals by flavonoids. Moreover, it appeared that when the free hydroxyl group was methoxylated or glycosylated, the inhibitory activity was obviously decreased or even abolished.

Antihypertensive substance in seeds of Areca catechu L.

Among various tannins tested, Areca II-5-C, a fraction isolated from seeds of Areca catechu L., showed the most potent angiotensin-converting enzyme (ACE) inhibitory activity in vitro. Its antihypertensive activity was therefore investigated in normotensive and spontaneous hypertensive rats (SHR) after both oral and intravenous (i.v.) administration. The activity was compared with that of captopril (D-3-mercapto-2-methylpropanoyl-L-proline), a potent ACE inhibitor. Oral administration of Areca II-5-C to SHR produced a lasting, dose-related antihypertensive effect, and the responses obtained with doses of 100 and 200 mg/kg were comparable to those of captopril at doses of 30 and 100 mg/kg. Intravenous administration of Areca II-5-C to SHR produced a rapid and marked reduction in blood pressure at doses of 10 and 15 mg/kg. The maximum antihypertensive effect of Areca II-5-C in SHR, at an i.v. dose of 15 mg/kg, was about 5 times as large as that of captopril at the same dose. Although the vasopressor response to norepinephrine and vasodepressor responses to bradykinin and acetylcholine were not appreciably changed by i.v. treatment with Areca II-5-C at a dose of 5 mg/kg, it did produce dose-related inhibition of the pressor responses to angiotensin I and II. It is suggested that Areca II-5-C has favorable properties as a hypotensive drug through its ability to inhibit the pressor responses to both angiotensin I and II.

Structure determination of polysaccharides in Aloe saponaria (Hill.) Haw. (Liliaceae).

Neutral polysaccharides that inhibit carrageenin-induced edema in rats were isolated from the nondialysate of the pulp of Aloe saponaria by gel filtration. These were shown to be a linear polymer of a 1,4-linked beta-D-mannopyranose (mol. wt. 15,000) containing 18% acetyl groups (As mannan 1), and a 1,4-linked alpha-D-mannopyranose polymer containing a single branch on the principal chain consisting of D-glucose residues linked at C-2 and C-4 (mol. wt. 66,000), with 10% acetyl groups (As mannan 2). As mannan 1 inhibited carrageenin-induced hind paw edema at 50 mg/kg ip in rats; As mannan 2 was not tested for pharmacological activity. A crude preparation of both As mannans was effective when given intraperitoneally, but was ineffective when given orally.

Antibradykinin active material in Aloe saponaria.

A material having antibradykinin activity on isolated guinea pig ileum was partially purified from the nondialysate of the pulp of Aloe saponaria by repetition of gel chromatography using a hydrophilic polyvinyl gel and dextran gels. From the results of amino acid and carbohydrate analyses, the antibradykinin-active material was estimated to be a glycoprotein. It was found that this material catalyzes the hydrolysis of bradykinin at pH 7.4. The results of peptide analysis using reversed-phase high-performance liquid chromatography coupled with amino acid analysis indicate that this glycoprotein cleaves the Gly4-Phe5 and Pro7-Phe8 bonds of the bradykinin molecule.

Tannins as potent inhibitors of DNA topoisomerase II in vitro.

Fifty-two out of 60 tannins, including gallo-, ellagi-, condensed, and complex tannins, are inhibitors of human DNA topoisomerase II in vitro. Thirty-six compounds that completely inhibited enzyme activity at a concentration of 500 nM or less, as assessed by ATP-dependent unknotting of P4 phage DNA, were at least 100-fold more potent than the clinically useful antitumor agent etoposide (VP-16). Relative inhibitory activity was primarily related to the number of phenolic hydroxyl groups (galloyl and hexahydroxydiphenoyl moieties) found in the active structures, with more groups generally conferring increased potencies. Unlike VP-16 and some DNA intercalative agents that stabilize the topoisomerase II-DNA cleavage intermediate, none of the active compounds induced protein-linked DNA breaks in cultured cells. Some of the tannins reduced VP-16-induced protein-linked DNA breaks by 20% or more, but one of these compounds, (-)-epicatechin, was not an inhibitor in vitro. Our data suggest that some tannins, such as sangiin H-6, that are potent inhibitors of catalytic double DNA-strand passage in vitro may target intracellular enzyme activity in a similar fashion to known poisons that interfere with formation of the enzyme-DNA covalent intermediate.

Renal responses to magnesium lithospermate B.

Renal responses to magnesium lithospermate B isolated from Salviae miltiorrhizae radix were examined in normal rats. Urinary sodium, potassium, prostaglandin E2 and kallikrein excretion was significantly increased after magnesium lithospermate B administration, whereas excretion of urinary 6-keto-prostaglandin F1 alpha and thromboxane B2 was unchanged. Rats administered with the drug also revealed a slight elevation of plasma renin activity and the levels of angiotensins I and II. Plasma aldosterone was decreased slightly. No significant changes were observed in angiotensin-converting enzyme or blood pressure.

Magnesium lithospermate B ameliorates cisplatin-induced injury in cultured renal epithelial cells.

A study was conducted to clarify whether magnesium lithospermate B ameliorates cisplatin-induced renal injury in terms of lactate dehydrogenase and malondialdehyde leakage from LLC-PK1 cells in culture. Magnesium lithospermate B was shown to suppress the cytotoxicity of cisplatin, the suppressive effect increasing with the dose of magnesium lithospermate B.

Isolation of a renal function-facilitating constituent from the Oriental drug, salviae miltiorrhizae radix.

An attempt was made to isolate the active component which exhibits an improving effect on renal function from Salviae Miltiorrhazae Radix (Chinese crude drug). Systematic isolation from aqueous extract of Salviae Miltiorrhizae Radix was carried out, and Compound 1 was found to be more effective than any of the other constituents in improving renal functional parameters; that is, a marked reduction of glomerular filtration rate following adenine ingestion was improved by administration of this substance. The renal plasma flow and renal blood flow were also increased in renal failure rats. On the basis of chemical and spectroscopic data, Compound 1 was shown to be identical with magnesium lithospermate B.

Effects of rhubarb tannins on renal function in rats with renal failure.

The effects of tannins purified from Rhei Rhizoma on various parameters of renal function were investigated in rats with adenine-induced renal failure. The glomerular filtration rate, renal plasma flow and renal blood flow were significantly increased in rats given (-)-epicatechin 3-O-gallate at a dose of 5 or 10 mg/kg body weight/day for 24 days. Administration of 5 mg of procyanidin B-2 3,3'-di-O-gallate also led to a significant increase in renal functional parameters. However, unlike the former two components, procyanidin C-1 3,3',3''-tri-O-gallate caused aggravation of renal function.

Magnesium lithospermate B improves renal function via the kallikrein-prostaglandin system in rats with renal failure.

The effect of magnesium lithospermate B on the renal responses of rats with renal failure was investigated in the presence and absence of pretreatment with the kallikrein inhibitor, aprotinin. Magnesium lithospermate B caused a marked increase in the levels of the renal functional parameters (glomerular filtration rate, renal plasma flow and renal blood flow), accompanied by significant increases in urinary prostaglandin excretion (increases of prostaglandin E2 and 6-keto-prostaglandin F1 alpha excretion by 82% and 36%, respectively). The urinary excretion of kallikrein was also increased following magnesium lithospermate B administration. However, pretreatment with aprotinin abolished the renal function-facilitating action of magnesium lithospermate B concomitantly with a markedly increased urinary excretion of prostaglandin E2, 6-keto-prostaglandin F1 alpha and kallikrein. These results suggest that the kallikrein-kinin-prostaglandin B.

Effect of magnesium lithospermate B on the renal and urinary kallikrein activities in rats with adenine-induced renal failure.

A study was conducted to examine the effect of magnesium lithospermate B on both the urinary and renal total and active kallikrein and prokallikrein in rats with adenine-induced renal failure. In rats given magnesium lithospermate B at a dose of 10 mg/kg body weight/day for 12 days, significant increases of urinary total and active kallikrein were associated with significant increases of urine volume and urinary total and active kallikrein were associated with significant increases of urine volume and urinary creatinine excretion. The renal total and active kallikrein levels were also significantly elevated by the treatment with magnesium lithospermate B. On day 24, the urinary excretion of total and active kallikrein and prokallikrein was significantly increased. Concomitantly, a significant increase in renal kallikrein (total, active and pro-) was found in the rats given magnesium lithospermate B. A significant relationship existed between the urinary creatinine and active kallikrein excretion. These results suggest that magnesium lithospermate B may stimulate the synthesis of kallikrein and/or conversion to active kallikrein, thus improving renal function.

Effects of a Dan Shen component, magnesium lithospermate B, in nephrectomized rats.

Magnesium lithospermate B, a compound newly isolated from Dan Shen, was given orally to rats for 70 days after excision of five-sixths of their kidney volume. As a result, mesangial proliferation, tubulo-interstitial lesions and glomerular sclerotic lesions, which were conspicuous in rats that were not given magnesium lithospermate B after nephrectomy, were inhibited. Furthermore, a decrease in blood urea nitrogen, improvement of hypoproteinemia, hypoalbuminemia and hypercholesteremia, and inhibition of urinary protein excretion were observed. The levels of creatinine, methylguanidine and guanidino-succinic acid, which accumulate in the blood with the progress of renal failure, were decreased significantly in rats given magnesium lithospermate B. These results indicate that magnesium lithospermate B, a component of an Oriental medicine has potential as a new therapeutic agent for inhibiting the progression of renal dysfunction.