Rational design and synthesis of topoisomerase I and II inhibitors based on oleanolic acid moiety for new anti-cancer drugs.

Semisynthetic reactions were conducted on oleanolic acid, a common plant-derived oleanane-type triterpene. Ten rationally designed derivatives of oleanolic acid were synthesized based on docking studies and tested for their topoisomerase I and IIα inhibitory activity. Semisynthetic reactions targeted C-3, C-12, C-13, and C-17. Nine of the synthesized compounds were identified as new compounds. The structures of these compounds were confirmed by spectroscopic methods (1D, 2D NMR and MS). Five oleanolic acid analogues (S2, S3, S5, S7 and S9) showed higher activity than camptothecin (CPT) in the topoisomerase I DNA relaxation assay. Four oleanolic acid analogues (S2, S3, S5 and S6) showed higher activity than etoposide in a topoisomerase II assay. The results indicated that the C12-C13 double bond of the oleanolic acid skeleton is important for the inhibitory activity against both types of topoisomerases, while insertion of a longer chain at either position 3 or 17 increases the activity against topoisomerases by various degrees. Some of the synthesized compounds act as dual inhibitors for both topoisomerase I and IIα.

Hypotensive effects and angiotensin-converting enzyme inhibitory peptides of reishi (Ganoderma lingzhi) auto-digested extract.

Reishi (Ganoderma lingzhi) has been used as a traditional medicine for millennia. However, relatively little is known about this mushroom's proteins and their bioactivities. In this study, we used reishi's own proteases to hydrolyze its protein and obtained auto-digested reishi (ADR) extract. The extract was subjected to in vitro assays and administered to spontaneous hypertensive rats (SHRs) to determine its potential for use as a hypotensive medication. Bioassay-guided fractionation and de novo sequencing were used for identifying the active compounds. After 4 h administration of ADR, the systolic pressure of SHRs significantly decreased to 34.3 mmHg (19.5% change) and the effect was maintained up to 8 h of administration, with the decrease reaching as low as 26.8 mmHg (15% reduction-compare to base line a decrease of 26.8 mmHg is less than a decrease of 34.3 mmHg so it should give a smaller % reduction). Eleven peptides were identified and four of them showed potent inhibition against ACE with IC50 values ranging from 73.1 µM to 162.7 µM. The results showed that ADR could be a good source of hypotensive peptides that could be used for antihypertensive medication or incorporation into functional foods.

Structure-activity relationships of lanostane-type triterpenoids from Ganoderma lingzhi as α-glucosidase inhibitors.

A series of lanostane-type triterpenoids, identified as ganoderma alcohols and ganoderma acids, were isolated from the fruiting body of Ganoderma lingzhi. Some of these compounds were confirmed as active inhibitors of the in vitro human recombinant aldose reductase. This paper aims to explain the structural requirement for α-glucosidase inhibition. Our structure-activity studies of ganoderma alcohols showed that the OH substituent at C-3 and the double-bond moiety at C-24 and C-25 are necessary to increase α-glucosidase inhibitory activity. The structure-activity relationships of ganoderma acids revealed that the OH substituent at C-11 is an important feature and that the carboxylic group in the side chain is essential for the recognition of α-glucosidase inhibitory activity. Moreover, the double-bond moiety at C-20 and C-22 in the side chain and the OH substituent at C-3 of ganoderma acids improve α-glucosidase inhibitory activity. These results provide an approach with which to consider the structural requirements of lanostane-type triterpenoids from G. lingzhi. An understanding of these requirements is considered necessary in order to improve a new type of α-glucosidase inhibitor.

Corncobs as a potential source of functional chemicals.

Phytochemical examination of corncob extracts led to the isolation of a new lignan identified as 7,7'-dihydroxy-3'-O-demethyl-4-methoxymatairesinol, together with seven known compounds, identified as ß-sitosterol, ß-sitosteryl-ß-D-glucoside, 6ß-hydroxy-campest-4-en-3-one, 5α,8α-epidioxyergosta-6,22-dien-3ß-ol, tricin, kaempferol and p-coumaric acid. The isolated compounds were identified by one and two-dimensional NMR spectroscopies and mass spectrometry.

Structure-activity relationships of ganoderma acids from Ganoderma lucidum as aldose reductase inhibitors.

A series of lanostane-type triterpenoids, known as ganoderma acids were isolated from the fruiting body of Ganoderma lucidum. Some of these compounds were identified as active inhibitors of the in vitro human recombinant aldose reductase. To clarify the structural requirement for inhibition, some structure-activity relationships were determined. Our structure-activity studies of ganoderma acids revealed that the OH substituent at C-11 is an important feature and the carboxylic group in the side chain is essential for the recognition of aldose reductase inhibitory activity. Moreover, double bond moiety at C-20 and C-22 in the side chain contributes to improving aldose reductase inhibitory activity. In the case of ganoderic acid C2, all of OH substituent at C-3, C-7 and C-15 is important for potent aldose reductase inhibition. These results provide an approach to understanding the structural requirements of ganoderma acids from G. lucidum for aldose reductase inhibitor. This understanding is necessary to design a new-type of aldose reductase inhibitor.

Cloning and transcriptional analysis of the gene encoding 5-aminolevulinic acid synthase of the white-rot fungus Phanerochaete sordida YK-624.

In this study, we cloned the gene encoding 5-aminolevulinic acid synthase (ALAS) from the hyper-lignin-degrading fungus Phanerochaete sordida YK-624. The deduced amino acid sequence showed highest identity (93.0%) to ALAS of P. chrysosporium. Expression of the gene encoding ALAS, which we named aas, corresponded temporally with the expression and activity of manganese peroxidase.

A novel metabolic pathway for biodegradation of DDT by the white rot fungi, Phlebia lindtneri and Phlebia brevispora.

1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT) was used as the substrate for a degradation experiment with the white rot fungi Phlebia lindtneri GB-1027 and Phlebia brevispora TMIC34596, which are capable of degrading polychlorinated dibenzo-p-dioxin (PCDD) and polychlorinated biphenyls (PCBs). Pure culture of P. lindtneri and P. brevispora with DDT (25 µmol l(-1)) showed that 70 and 30% of DDT, respectively, disappeared in a low-nitrogen medium after a 21-day incubation period. The metabolites were analyzed using gas chromatography/mass spectrometry (GC/MS). Both fungi metabolized DDT to 1,1-dichloro-2,2-bis(4-chlorophenyl)ethane (DDD), 2,2-bis(4-chlorophenyl)acetic acid (DDA) and 4,4-dichlorobenzophenone (DBP). Additionally, DDD was converted to DDA and DBP. DDA was converted to DBP and 4,4-dichlorobenzhydrol (DBH). While DBP was treated as substrate, DBH and three hydroxylated metabolites, including one dihydroxylated DBP and two different isomers of monohydroxylated DBH, were produced from fungal cultures, and these hydroxylated metabolites were efficiently inhibited by the addition of a cytochrome P-450 inhibitor, piperonyl butoxide. These results indicate that the white rot fungi P. lindtneri and P. brevispora can degrade DBP/DBH through hydroxylation of the aromatic ring. Moreover, the single-ring aromatic metabolites, such as 4-chlorobenzaldehyde, 4-chlorobenzyl alcohol and 4-chlorobenzoic acid, were found as metabolic products of all substrate, demonstrating that the cleavage reaction of the aliphatic-aryl carbon bond occurs in the biodegradation process of DDT by white rot fungi.

Melanin biosynthesis inhibitory and antioxidant activities of quercetin-3'-O-beta-D-glucoside isolated from Allium cepa.

In the course of searching for new whitening agents, we have found that the methanol extract of dried skin of Allium cepa shows potent melanin biosynthesis inhibitory activity in B16 melanoma cells. Bioassay-guided fractionation led to the isolation of quercetin-3'-O-beta-D-glucoside (1) from the methanol extract of dried skin of A. cepa, which inhibited melanin formation in B16 melanoma cells with an IC50 value of 38.8 microM and mushroom tyrosinase with an IC50 value of 6.5 microM using L-tyrosine and 48.5 microM using L-dihydroxyphenylalanine as substrates, respectively. In addition, the antioxidant activity of 1 was evaluated in the oxygen radical absorbance capacity assay; it showed 3.04 micromol Trolox equivalents/mmol. 1 was shown to be a promising ingredient that could be useful for treating hyperpigmentation and for protecting against oxidative stress.

Ganoderic acids from Ganoderma lucidum: inhibitory activity of osteoclastic differentiation and structural criteria.

Increased osteoclastic bone resorption plays a central role in the pathogenesis of many bone diseases. Ganoderma lucidum is known for medicinal effects such as anti-inflammatory and anti-bone resorption activities. In this study, we investigated the inhibitory effects of ganoderic acids isolated from G. lucidum on osteoclastic differentiation using RAW264 cells in vitro. A carbonyl in C7 is essential to elicit the inhibition of osteoclast differentiation.

Inhibition of aldose reductase in vitro by constituents of Ganoderma lucidum.

CHCl(3) extract of the fruiting body of Ganoderma lucidum was found to show inhibitory activity on human aldose reductase in vitro. From the acidic fraction, potent human aldose reductase inhibitors, ganoderic acid C2 (1) and ganoderenic acid A (2), were isolated together with three related compounds. It was found that the free carboxyl group of ganoderic acid C2 and ganoderenic acid A is essential in eliciting the inhibitory activity considering the much lower activity of their methyl esters.

Potency of Phlebia species of white rot fungi for the aerobic degradation, transformation and mineralization of lindane.

The widespread use of the organochlorine insecticide lindane in the world has caused serious environmental problems. The main purpose of this paper is to investigate the potency of several Phlebia species of white rot fungi to degrade, transform and mineralize lindane, and to provide the feasibility of using white rot fungi for bioremediation at contaminated sites. Based on tolerance experiment results, Phlebia brevispora and Phlebia lindtneri had the highest tolerance to lindane and were screened by degradation tests. After 25 days of incubation, P. brevispora and P. lindtneri degraded 87.2 and 73.3% of lindane in low nitrogen medium and 75.8 and 64.9% of lindane in high nitrogen medium, respectively. Several unreported hydroxylation metabolites, including monohydroxylated, dehydroxylated, and trihydroxylated products, were detected and identified by GC/MS as metabolites of lindane. More than 10% of [14C] lindane was mineralized to 14CO2 by two fungi after 60 days of incubation, and the mineralization was slightly promoted by the addition of glucose. Additionally, the degradation of lindane and the formation of metabolites were efficiently inhibited by piperonyl butoxide, demonstrating that cytochrome P450 enzymes are involved in the fungal transformation of lindane. The present study showed that P. brevispora and P. lindtneri were efficient degraders of lindane; hence, they can be applied in the bioremediation process of lindane-contaminated sites.

20(R)-ginsenoside Rh2, not 20(S), is a selective osteoclastgenesis inhibitor without any cytotoxicity.

Increased osteoclastic bone resorption plays a central role in the pathogenesis of many bone diseases, and osteoclast inhibitors are the most widely used treatments for these diseases. Ginsenosides, the main component of ginseng, have been known for their medicinal effects such as anti-inflammatory and anti-proliferative activities. In this study, we investigated the inhibitory effects of ginsenosides (ginsenoside 20(R)-Rh2 and ginsenoside 20(S)-Rh2) on osteoclastgenesis using RAW264 cells in vitro. Only ginsenoside 20(R)-Rh2 showed selective osteoclastgenesis inhibitory activity without any cytotoxicity up to 100 microM. These results implied that the stereochemistry of the hydroxyl group at C-20 may play an important role in selective osteoclastgenesis inhibitory activity.

Ganoderic acid DM: anti-androgenic osteoclastogenesis inhibitor.

Prostate cancer is the most common cancer in men in Western countries, with a high incidence of bone metastasis. Ganoderic acid DM, with 5alpha-reductase inhibitory and androgen receptor (AR) binding activity, isolated from the ethanol extracts of Ganoderma lucidum, can inhibit prostate cancer cell growth and block osteoclastogenesis.

The inhibitory effect on aldose reductase by an extract of Ganoderma lucidum.

The human aldose reductase inhibitory effects of the methanol extracts of 17 medicinal and edible mushrooms were examined. Ganoderma lucidum showed the highest aldose reductase inhibitory activity compared with the other mushrooms. The effect of an ethanol extract of G. lucidum on the galactitol level in the eye lens was studied in a galactosemic rat model in vivo. This mushroom significantly decreased the galactitol accumulation.

Anti-androgenic activity of fatty acids.

In this study, we show that 5alpha-reductase derived from rat fresh liver was inhibited by certain aliphatic free fatty acids. The influences of chain length, unsaturation, oxidation, and esterification on the potency to inhibit 5alpha-reductase activity were studied. Among the fatty acids we tested, inhibitory saturated fatty acids had C12-C16 chains, and the presence of a C==C bond enhanced the inhibitory activity. Esterification and hydroxy compounds were totally inactive. Finally, we tested the prostate cancer cell proliferation effect of free fatty acids. In keeping with the results of the 5alpha-reductase assay, saturated fatty acids with a C12 chain (lauric acid) and unsaturated fatty acids (oleic acid and alpha-linolenic acid) showed a proliferation inhibitory effect on lymph-node carcinoma of the prostate (LNCaP) cells. At the same time, the testosterone-induced prostate-specific antigen (PSA) mRNA expression was down-regulated. These results suggested that fatty acids with 5alpha-reductase inhibitory activity block the conversion of testosterone to 5alpha-dihydrotestosterone (DHT) and then inhibit the proliferation of prostate cancer cells.

Anti-androgen effects of extracts and compounds from Ganoderma lucidum.

The 30% EtOH extracts of Ganoderma lucidum Fr. Karst (Ganodermateceae) showed weak 5alpha-reductase inhibitory activity and binding ability to androgen receptor. When LNCaP (lymph-node carcinoma of the prostate) cells were treated with the EtOH extracts, cell proliferation was inhibited. Treatment with the extracts significantly inhibited the testosterone-induced growth of the ventral prostate in castrated rats. These results showed that G. lucidum might be a useful ingredient in the treatment of androgen-induced diseases, such as benign prostatic hyperplasia or prostate cancer. From the 30% EtOH extracts, we isolated ganoderiol F, which showed binding activity to androgen receptor and inhibited LNCaP cell proliferation, as one of the active compounds in the 30% EtOH extracts.

Saline-dependent regulation of manganese peroxidase genes in the hypersaline-tolerant white rot fungus Phlebia sp. strain MG-60.

The expression pattern of manganese peroxidases (MnPs) in nitrogen-limited cultures of the saline-tolerant fungus Phlebia sp. strain MG-60 is differentially regulated under hypersaline conditions at the mRNA level. When MG-60 was cultured in nitrogen-limited medium (LNM) containing 3% (wt/vol) sea salts (LN-SSM), higher activity of MnPs was observed than that observed in normal medium (LNM). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis demonstrated that two MnP isoenzymes were de novo synthesized in the culture of LN-SSM. Three MnP-encoding genes (MGmnp1, MGmnp2, and MGmnp3) were isolated by reverse transcription (RT)-PCR and rapid amplification of cDNA ends PCR techniques. The corresponding isozymes were identified by peptide mass fingerprinting analysis. MnP isozymes encoded by MGmnp2 and MGmnp3 were observed mainly in LN-SSM. Real-time RT-PCR analysis revealed high levels of MGmnp2 and MGmnp3 transcripts in LN-SSM 48 h after the addition of 2% NaCl. The induction of MnP production and the accumulation of gene transcripts by saline were well correlated in the presence of Mn(2+). However, in the absence of Mn(2+), there was no clear correlation between mnp transcripts levels and MnP activity, suggesting posttranscriptional regulation by Mn(2+).

Role of Tyr residues on the protein surface of cationic cell-wall-peroxidase (CWPO-C) from poplar: potential oxidation sites for oxidative polymerization of lignin.

It was previously reported that an unique peroxidase isoenzyme, cationic cell-wall-bound peroxidase (CWPO-C), from poplar callus oxidizes sinapyl alcohol, ferrocytochrome c and synthetic lignin polymers, unlike other plant peroxidases. Here, the catalytic mechanism of CWPO-C was investigated using chemical modification and homology modeling. The simulated CWPO-C structure predicts that the entrance to the heme pocket of CWPO-C is the same size as those of other plant peroxidases, suggesting that ferrocytochrome c and synthetic lignin polymers cannot interact with the heme of CWPO-C. Since Trp and Tyr residues are redox-active, such residues located on the protein surface were predicted to be active sites for CWPO-C. Modification of CWPO-C Trp residues did not suppress its oxidation activities toward guaiacol and syringaldazine. On the other hand, modification of CWPO-C Tyr residues using tetranitromethane strongly suppressed its oxidation activities toward syringaldazine and 2,6-dimethoxyphenol by 90%, respectively, and also suppressed its guaiacol oxidation activity to a lesser extent. Ferrocytochrome c was not oxidized by Tyr-modified CWPO-C. These results indicate that the Tyr residues in CWPO-C mediate its oxidation of syringyl compounds and high-molecular-weight substrates. Homology modeling indicates that Tyr-177 and Tyr-74 are located near the heme and exposed on the protein surface of CWPO-C. These results suggest that Tyr residues on the protein surface are considered to be important for the oxidation activities of CWPO-C with a wide range of substrates, and potentially unique oxidation sites for the plant peroxidase family.

Essential oil of lavender inhibited the decreased attention during a long-term task in humans.

This study examined the effects of odors on sustained attention during a vigilance task. Two essential oils (lavender and eucalyptus) and two materials (l-menthol and linalyl acetate) were compared with a control. The increase in reaction time was significantly lower with lavender than with the control. The results suggest that the administration of lavender helped to maintain sustained attention during the long-term task.

Effect of an extract of Ganoderma lucidum in men with lower urinary tract symptoms: a double-blind, placebo-controlled randomized and dose-ranging study.

AIM: To conduct a double-blind, placebo-controlled randomized and dose-ranging study to evaluate the safety and efficacy of the extract of Ganoderma lucidum (G. lucidum) in men with lower urinary tract symptoms (LUTS). METHODS: We enrolled male volunteers (> or = 50 years) with an International Prostate Symptom Score (IPSS; questions 1-7) > or = 5 and a prostate-specific antigen (PSA) value < 4 ng/mL. Volunteers were randomized into groups of placebo (n = 12), G. lucidum of 0.6 mg (n = 12), 6 mg (n = 12) or 60 mg (n = 14), administered once daily. Efficacy was measured as a change from baseline in IPSS and the peak urine flow rate (Q(max)). Prostate volume and residual urine were estimated by ultrasonography, and blood tests, including PSA levels, were measured at baseline and at the end of the treatment. RESULTS: The overall administration was well tolerated, with no major adverse effects. Statistical significances in the magnitude of changes between the experimental groups were observed at weeks 4 and 8. No changes were observed with respect to Q(max), residual urine, prostate volume or PSA levels. CONCLUSION: The extract of G. lucidum was well tolerated and an improvement in IPSS was observed. The recommended dose of the extract of G. lucidum is 6 mg in men with LUTS.