Rat lens aldose reductase inhibitory activities of Coptis japonica root-derived isoquinoline alkaloids

The inhibitory activity of Coptis japonica root-derived materials was evaluated against lens aldose reductase isolated from male Sprague-Dawley rats and compared to that of three commercially available isoquinoline alkaloids (berberine sulfate, berberine iodide, and palmatine chloride), as well as quercitrin as aldose reductase inhibitor. The biologically active constituents of C. japonica extract were characterized as the isoquinoline alkaloids, berberine chloride and palmatine iodide, by spectral analysis. The inhibitory effects varied with both chemical and concentration used. The IC(50) values of berberine chloride and palmatine iodide are 13.98 and 13.45 nM, respectively. Among three berberines and two palmatines, the inhibitory activity was much greater for the choridated and sulfated analogues than for those with iodide. Quercitrin was a much more potent inhibitor than berberines and palmatines. Nonetheless, berberines and palmatines may be useful as lead compounds and new agents for aldose reductase inhibition.     

Growth-inhibiting effects of Coptis japonica root-derived isoquinoline alkaloids on human intestinal bacteria.

The growth-inhibiting activity of Coptis japonica (Makino) root-derived materials toward eight human intestinal bacteria was examined using an impregnated paper disk method and compared to that of four commercially available isoquinoline alkaloids [berberine sulfate (BS), berberine iodide (BI), palmatine chloride (PC), and palmatine sulfate(PS)], as well as that of Thea sinensis leaf-derived epigallocatechin gallate (EGCG). The biologically active constituents of the Coptis extract were characterized as the isoquinoline alkaloids berberine chloride (BC), palmatine iodide (PI), and coptisine chloride (CC) by spectral analysis. The growth responses varied with both chemical and bacterial strain used. In a test using 500 microg/disk, BC and PI produced a clear inhibitory effect against Bifidobacterium longum, Bifidobacterium bifidum, Clostridium perfringens, and Clostridium paraputrificum, whereas weak or no inhibition was observed in Bifidobacterium adolescentis, Lactobacillus acidophilus, Lactobacillus casei, and Escherichia coli. At 1000 microg/ disk, CC revealed weak or no growth inhibition toward all test bacteria, whereas EGCG exhibited weak growth inhibition against only C. perfringens and C. paraputrificum. Among various isoquinoline alkaloids, BC exhibited more potent inhibitory activity toward C. perfringens than BI and BS, whereas the inhibitory effect was more pronounced in PI compared to PC and PS. The Coptis root-derived materials did not promote growth of B. longum and C. perfringens.     

Antioxidant activity of phenolic compounds isolated from Mesona procumbens Hemsl

The antioxidant activity of phenolic compounds isolated from Mesona procumbens Hemsl. (Hsian-tsao) was investigated. Hsian-tsao was extracted with various solvents, and the results showed that the fraction treated with acidic ethyl acetate (pH 2) possessed large amounts of phenolic compounds and a strong antioxidant activity on peroxidation of linoleic acid. The antioxidant activity (inhibition of peroxidation, IP%) of the acidic ethyl acetate of Hsian-tsao extract at 50 microg/mL (98.9%) was stronger than those of 50 microg/mL alpha-tocopherol (78%) and BHA at 10 microg/mL (90%). When fractionated with Amberlite XAD-7 gel chromatography, the acidic ethyl acetate fraction of Hsian-tsao extract was separated into four subfractions (A-D). Subfraction B, with high yield and strong antioxidant activity, was further isolated and purified and then identified as containing protocatechuic acid, p-hydroxybenzoic acid, vanillic acid, caffeic acid, and syringic acid by means of UV, EI-MS, and (1)H and (13)C NMR. The antioxidant capability of isolated compounds was also determined using the thiocyanate system and the erythrocyte ghost system. The results indicate that the phenolic acids could be important antioxidant components in Hsian-tsao, among which caffeic acid with the highest antioxidant activity and the greatest content is most important.     

Identification of an antioxidant,ethyl protocatechuate,in peanut seed testa

The antioxidant activity and identification of the antioxidant component of peanut seed testa were investigated. The antioxidant activity of peanut seed testa was studied in the linoleic acid model system by using the ferric thiocyanate method. Among the five organic solvent extracts, the ethanolic extracts of peanut seed testa (EEPST) produced higher yields and stronger antioxidant activity than other organic solvent extracts. EEPST was separated into 17 fractions on silica gel column chromatography. Fraction 17, which showed the largest yield and significant antioxidant activity, was separated by thin-layer chromatography. Four major antioxidative subfractions were present. Subfraction 17-2 was found to be effective in preventing oxidation of linoleic acid. This subfraction was further fractionated and isolated and characterized by UV, MS, IR, and (1)H NMR techniques. The active compound was identified as ethyl protocatechuate (3,4-dihydroxybenzoic acid ethyl ester).     

Isolation and characterization of an oxygen radical absorbance activity peptide from defatted peanut meal hydrolysate and its antioxidant properties

Defatted peanut meal hydrolysate (DPMH) was purified using ultrafiltration, gel filtration chromatography, and high-performance liquid chromatography. A tripeptide with strong oxygen radical absorbance capacity (ORAC) was isolated and identified as Tyr-Gly-Ser by ESI-MS/MS. It was then synthesized to measure its antioxidant properties in different systems. The ORAC value of Tyr-Gly-Ser was 3-fold higher than that of glutathione (GSH), and it displayed a stronger protective effect on linoleic acid peroxidation and H(2)O(2)-induced oxidative injury in rat pheochromocytoma line PC12 cells than GSH (p < 0.05). However, Tyr-Gly-Ser showed negligible DPPH radical scavenging activity, reducing power, and no metal chelating ability. The results suggested that Tyr-Gly-Ser displayed antioxidant activity via the hydrogen atom transfer mechanism, and the Tyr at the N-terminal was the hydrogen donor. The ORAC assay was recommended as a reliable and effective method to measure the antioxidant activity in the course of antioxidant peptide isolation.     

Antioxidative properties of tripeptide libraries prepared by the combinatorial chemistry

Two series of combinatorial tripeptide libraries were constructed, based on an antioxidative peptide isolated from a soybean protein hydrolysate. One was a library of 108 peptides containing either His or Tyr residues. Another was a library of 114 peptides related to Pro-His-His, which had been identified as an active core of the antioxidative peptide. The antioxidative properties of these libraries were examined by several methods, such as the antioxidative activity against the peroxidation of linoleic acid, the reducing activity, the radical scavenging activity, and the peroxynitrite scavenging activity. Two Tyr-containg tripeptides showed higher activities than those of two His-containing tripeptides in the peroxidation of linoleic acid. Tyr-His-Tyr showed a strong synergistic effects with phenolic antioxidants. However, the tripeptide had only marginal reducing activity and a moderate peroxynitrite scavenging activity. Cysteine-containing tripeptides showed the strong peroxynitrite scavenging activity. Change of either the N-terminus or C-terminus of Pro-His-His to other amino acid residues did not significantly alter their antioxidative activity. Tripeptides containing Trp or Tyr residues at the C-terminus had strong radical scavenging activities, but very weak peroxynitrite scavenging activity. The present results allow us to understand why protein digests have such a variety of antioxidative properties.     

Evaluation of the antioxidant activity of environmental plants: activity of the leaf extracts from seashore plants

The antioxidant activity of the methanolic extracts of the leaves of 39 plant species was examined. These leaves were collected from the plants growing on subtropical seashores. The activity was evaluated by three kinds of assay methods, which included the DPPH radical scavenging assay, linoleic acid oxidation assay, and oxidative cell death assay. Two extracts from Excoecaria agallocha and Terminalia catappa showed remarkably potent activity in all assay systems. The HPLC analysis of the extracts indicated the presence of the same antioxidant and isolation work for the compound identified ellagic acid. The isolated ellagic acid showed strong antioxidant activity in the assay systems used.     

Evaluation of antioxidant activity of vetiver (Vetiveria zizanioides L.) oil and identification of its antioxidant constituents

Antioxidant capacities of vetiver (Vetiveria zizanioides) oil were evaluated by two different in vitro assays: the DPPH* free radical scavenging assay and the Fe2+-metal chelating assay. Results showed that the vetiver oil (VO) possessed a strong free radical scavenging activity when compared to standard antioxidants such as butylated hydroxytoluene (BHT) and alpha-tocopherol. However, its metal chelating capacity was relatively weak. VO (10 microL/mL) dissolved in methanol exhibited approximately 93% free radical scavenging activity in the DPPH* assay and approximately 34% Fe2+ chelating activity in the metal chelating assay. By contrast, 10 mM BHT and 0.1 mM alpha-tocopherol exhibited 93 and 89% free radical scavenging activities in the DPPH* assay, respectively, and 1 mM EDTA exhibited approximately 97% activity in the metal chelating assay. Among the complex constituents in the crude VO, beta-vetivenene, beta-vetivone, and alpha-vetivone, which had shown strong antioxidant activities, were isolated and identified using various chromatographic techniques including silica gel open column chromatography, silica HPLC, and GC-MS. These results show that VO and some of its inherent components can be potential alternative natural antioxidants.     

Cordycepin: selective growth inhibitor derived from liquid culture of Cordyceps militaris against Clostridium spp

The growth responses of nine human intestinal bacteria to liquid culture of Cordyceps militaris Link. Pt. (Ascomycotina: Clavicipitaceae) collected from a pupa of Bombyx mori L. (Lepidoptera: Bombycidae) were examined using spectrophotometric and impregnated paper disk methods and compared to those of tetracycline and chloramphenicol, as well as those of Coptis japonica root-derived berberine chloride. The biologically active constituent of the cultures was characterized as cordycepin (3'-deoxyadenosine) by spectroscopic analysis. This compound revealed potent growth-inhibiting activity toward Clostridium paraputrificum and Clostridium perfringens at 10 microgram/disk without adverse effects on the growth of Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium longum, Bifidobacterium adolescentis, Lactobacillus acidophilus, and Lactobacillus casei, whereas tetracycline and chloramphenicol inhibited the growth of these lactic acid-producing bacteria, clostridia and Escherichia coli. However, C. militaris-derived materials revealed no growth stimulation on the bifidobacteria and lactobacilli. These results may be an indication of at least one of the pharmacological actions of C. militaris. As a naturally occurring antibacterial agent, cordycepin could be useful as a new preventive agent against various diseases caused by clostridia.     

Chemical composition and antioxidant activity of phenolic compounds from wild and cultivated Sclerocarya birrea(Anacardiaceae) leaves

A quantitative study of the phenolic constituents of wild and cultivated leaves of Sclerocarya birrea(Anacardiaceae) was carried out by HPLC-UV/PDA and LC-MS. Phytochemical analysis of the methanol extract of wild plants led to the isolation of one new flavonol glycoside, quercetin 3-O-alpha-l-(5' '-galloyl)-arabinofuranoside (1), and eight known phenolic compounds; two epicatechin derivatives were also isolated from the same extract of the cultivated species. The antioxidant activity of all isolated compounds was determined by measuring free radical scavenging effects using the Trolox equivalent antioxidant capacity assay and the coupled oxidation of beta-carotene and linoleic acid (autoxidation assay).     

Antioxidative activity in the pericarp and seed of Japanese pepper (Xanthoxylum piperitum DC)

The antioxidative activity of the dried pericarp and seed of Japanese pepper was studied. The ethyl acetate extract from the pericarp and the methanol extract from the seed showed strong antioxidative activity against linoleic acid by the ferric thiocyanate and thiobarbituric acid (TBA) methods. Japanese pepper contained 3.9 and 2.9 mg/100 g of dry weight (dw) of tocopherols in the pericarp and seed, respectively, alpha-Toc in the former constituting 82% of total tocopherol and gamma-Toc in the latter constituting 96%. Arbutin and magnoflorine were isolated as antioxidants and their chemical structures determined by instrumental analyses. The contents of arbutin evaluated as the trifluoroacetate derivative by GC-MS were 35 and 3.0 mg/100 g of dw in the pericarp and seed, respectively. Magnoflorine was present only in the seed, and not in the pericarp. Both arbutin and magnoflorine exhibited antioxidative activity against linoleic acid and radical-scavenging activity against the DPPH radical.     

Antiphoto-oxidative activity of sesamol in methylene blue- and chlorophyll-sensitized photo-oxidation of oil

The effects and mechanism of sesamol on the methylene blue- or chlorophyll-sensitized photo-oxidations of soybean oil have been studied. Sesamol showed strong antiphoto-oxidative activity in both methylene blue-and chlorophyll-sensitized photo-oxidations of soybean oil in a dose-dependent manner. The 1.0 x 10(-3) M sesamol treatments showed 84.7 and 43.4% inhibitions of methylene blue- and chlorophyll-sensitized photo-oxidations of soybean oil in methylene chloride. The antiphoto-oxidative activity of sesamol was comparable to that of delta-tocopherol in both methylene blue- and chlorophyll-sensitized photo-oxidations, at the same molar basis. Sesamol effectively inhibited rubrene oxidation with a chemical source of singlet oxygen in microemulsion, showing its strong singlet oxygen quenching ability. The results suggested that the antiphoto-oxidative activity of sesamol in the photo-oxidation of oil was, at least in part, due to its singlet oxygen scavenging activity. The singlet oxygen quenching rate constant (k(ox-Q) + k(q)) of sesamol was determined to be 1.9 +/- 0.3 x 10(7) M(-1) s(-1). This represents the first report on the antiphoto-oxidative activity of sesamol in the sensitized photo-oxidation of oil, and its bimolecular singlet oxygen quenching ability.     

Antioxidant properties of ferulic acid and its related compounds

Antioxidant activity of 24 ferulic acid related compounds together with 6 gallic acid related compounds was evaluated using several different physical systems as well as their radical scavenging activity. The radical scavenging activity on 1,1-diphenyl-2-picrylhydrazyl (DPPH) decreased in the order caffeic acid > sinapic acid > ferulic acid > ferulic acid esters > p-coumaric acid. In bulk methyl linoleate, test hydroxycinnamic acids and ferulic acid esters showed antioxidant activity in parallel with their radical scavenging activity. In an ethanol-buffer solution of linoleic acid, the activity of test compounds was not always associated with their radical scavenging activity. Ferulic acid was most effective among the tested phenolic acids. Esterification of ferulic acid resulted in increasing activity. The activity of alkyl ferulates was somewhat influenced by the chain length of alcohol moiety. When the inhibitory effects of alkyl ferulates against oxidation of liposome induced by AAPH were tested, hexyl, octyl, and 2-ethyl-1-hexyl ferulates were more active than the other alkyl ferulates. Furthermore, lauryl gallate is most effective among the tested alkyl gallates. These results indicated that not only the radical scavenging activity of antioxidants, but also their affinity with lipid substrates, might be important factors in their activity.     

Studies on the constituents of Cyclanthera pedata fruits: isolation and structure elucidation of new flavonoid glycosides and their antioxidant activity.

The isolation of six flavon glycosides (1-6), among them four new natural compounds (1-4), from the CHCl(3)/MeOH extract of the fruits of Cyclanthera pedata is reported. All of the structures were elucidated by spectroscopic methods, including the concerted application of one-dimensional ((1)H, (1)H TOCSY, (13)C, and (13)C DEPT-NMR) and two-dimensional NMR techniques (DQF-COSY, HSQC, and HMBC). For all of the isolated compounds the antioxidant activity was determined by measuring the free radical scavenging activity, using the Trolox equivalent antioxidant capacity (TEAC) method, and the coupled oxidation of beta-carotene and linoleic acid.     

Isolation of antioxidant compounds from the methanolic extract of the roots of Decalepis hamiltonii (Wight and Arn.)

The tuberous roots of Decalepis hamiltonii are consumed as pickles and beverages and are believed to possess health-promoting properties. We have investigated the antioxidant potential of the roots. The methanolic extract of the root showed a high antioxidant activity. The methanolic extract was fractionated on a silica gel column, which showed three major fractions with good antioxidant activity. The active fractions were further subjected to preparative thin layer and silica gel column chromatography, which yielded six pure compounds. The purified compounds were characterized by MS, 1H NMR, 13C NMR, and two-dimensional NMR spectroscopic techniques and identified as 2-hydroxy-4-methoxybenzaldehyde, p-anisaldehyde, vanillin, borneol, salicylaldehyde, and bis-2,3,4,6-galloyl-alpha/beta-D-glucopyranoside. The latter compound, named decalepin, is a new antioxidant molecule from the plant kingdom. The purified compounds showed antioxidant activities in in vitro assays such as inhibition of lipid peroxidation, hydroxyl radical, superoxide anion, and 1,1-diphenyl-2picrylhydrazyl radical scavenging. This is the first report of the antioxidant constituents of the roots of Decalepis hamiltonii.     

Free radical scavenging active components from Cedrus deodara

An activity-directed fractionation and purification process was used to identify the antioxidant components of Cedrus deodara. Dried heartwood powder of C. deodara was first defatted with petroleum ether and then extracted with chloroform. The chloroform extract showed strong antioxidant activity on 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical. This fraction was then subjected to separation and purification using silica gel column chromatography. Three compounds with potent antioxidant activity were isolated in significant yields and identified by spectroscopic methods ((1)H NMR, (13)C NMR, IR, and MS). They were identified as (-)-matairesinol, (-)-nortrachelogenin, and a dibenzylbutyrolactollignan (4,4',9-trihydroxy-3,3'-dimethoxy-9,9'-epoxylignan). This is the first report of the occurrence of these compounds in C. deodara.     

Free radical scavenging and antioxidative activity of caffeic acid amide and ester analogues: structure-activity relationship.

The structure-activity relationships of synthetic caffeic acid amide and ester analogues as potential antioxidants and free radical scavengers have been investigated. The 2,2-diphenyl-1-picrylhydrazyl radical (DPPH.) scavenging activity of the test compounds was N-trans-caffeoyl-L-cysteine methyl ester (5) > N-trans-caffeoyldopamine (4) > N-trans-caffeoyltyramine (3) > N-trans-caffeoyl-beta-phenethylamine (2) > Trolox C (8) > caffeic acid phenethyl ester (1) > caffeic acid (6) > ferulic acid (7). This established that the radical scavenging activity of the compounds increased with increasing numbers of hydroxyl groups or catechol moieties and also with the presence of other hydrogen-donating groups (-NH, -SH). The antioxidative activity of the compounds was also investigated in an emulsified linoleic acid oxidation system accelerated by 2,2'-azobis(2-amidinopropane) dihydrochloride. The order was 1 > 2 > 4 > 3 > or = 5 > 6 > 8 > 7. Therefore, in the emulsion system, the antioxidative activity of the test compounds depends not only on the hydroxyl groups or catechol rings but also on the partition coefficient (log P) or hydrophobicity of the compounds. This supports the concept that hydrophobic antioxidants tend to exhibit better antioxidative activity in an emulsion system.     

Phytotoxic Eremophilanes from Ligularia macrophylla

Systematic bioassay-guided fractionation of the methylene chloride extract of the roots from Ligularia macrophylla was performed to identify both phytotoxic and antifungal compounds. Four phytotoxic eremophilanes (furanoeremophilan-14beta,6alpha-olide, 6beta-angeloyloxy-10beta-hydroxyfuranoeremophilane, eremophil-7(11)-ene-12,8alpha;14beta,6alpha-diolide, and 3alpha-angeloyloxybakkenolide A) and two antifungal fatty acids (linoleic acid and alpha-linolenic acid) were isolated. The X-ray crystal structure determination of 6beta-angeloyloxy-10beta-hydroxyfuranoeremophilane is reported here for the first time. All four eremophilanes substantially inhibited growth of the monocot Agrostis stolonifera (bentgrass) while demonstrating little activity against the dicot Lactuca sativa (lettuce) at 1000 microM. In a dose-response screening of all compounds for growth inhibitory activity against Lemna paucicostata, 6beta-angeloyloxy-10beta-hydroxyfuranoeremophilane was the most active with an IC50 of 2.94+/-0.16 microM. This compound also caused the greatest reduction of photosynthetic electron flow; however, its mode of action remains to be determined. Evaluation of isolated compounds for activity against the Formosan subterranean termite, Coptotermes formosanus, is also reported. At a concentration of 0.5% (wt/wt), 6beta-angeloyloxy-10beta-hydroxyfuranoeremophilane significantly reduced the consumption of filter paper by C. formosanus.     

Synergistic effects of thiols and amines on antiradical efficiency of protocatechuic acid

DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging activity of protocatechuic acid and its structural analogues (methyl protocatechuate, 3',4'-dihydroxyacetophenone, 3,4-dihydroxybenzaldehyde, and 3,4-dihydroxybenzonitrile) were examined in aprotic and protic solvents. In aprotic acetonitrile, all test compounds scavenged two radicals. In protic methanol, however, these compounds rapidly scavenged five radicals except for protocatechuic acid, which consumed only two radicals. The result indicated that higher radical scavenging activity in methanol than in acetonitrile was due to a nucleophilic addition of the methanol molecule on the oxidized quinones, which led to a regeneration of catechol structures. To investigate the importance of the nucleophilic addition on the quinones for the high radical scavenging activity, DPPH radical scavenging activity of protocatechuic acid and its analogues was examined in the presence of a variety of nucleophiles. The addition of a strong nucleophile such as a cysteine derivative significantly increased the radical scavenging equivalence. Furthermore, thiol adducts at C-2 and C-2,5 of protocatechuic acid and its analogues were isolated from the reaction mixtures. These results strongly suggest that the quinone of protocatechuic acid and its analogues undergo a nucleophilic attack at C-2 to yield 2-substituted-3,4-diols. Then, a regenerated catechol moiety of adducts scavenge two additional radicals by reoxidation into quinones, which undergo the second nucleophilic attack at the C-5. This mechanism demonstrates a possibility of synergistic effects of various nucleophiles on the radical scavenging ability of plant polyphenols containing a 3,4-dihydroxy substructure like protocatechuic acid and its analogues.     

Identification of six phenylpropanoids from garlic skin as major antioxidants

The extract of garlic skins (peels) showed strong antioxidant activity, and some responsible constituents were isolated and identified. Garlic (Allium sativum L.) has been used as an herbal medicine, but there is no report on the health benefits of the skin or peel. In this study, the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity of garlic skin extract was evaluated. Using chromatographic techniques, the active constituents were isolated and subsequently identified. Analyses by high-performance liquid chromatography coupled with a photodiode array detector (HPLC-PDA) suggested that these compounds were phenylpropanoids, which had a characteristic absorbance at 300-320 nm. Liquid chromatography-mass spectrometry (LC-MS) and nuclear magnetic resonance analyses allowed the chemical structures of the isolated constituents to be postulated. The proposed compounds were subsequently synthesized and compared with the constituents in the extract using HPLC-PDA and LC-MS. N-trans-Coumaroyloctopamine, N-trans-feruloyloctopamine, guaiacylglycerol-beta-ferulic acid ether, and guaiacylglycerol-beta-caffeic acid ether were identified as were trans-coumaric acid and trans-ferulic acid. Also, the antioxidant activities of these compounds were determined.