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.     

Novel low-density lipoprotein (LDL) oxidation model: antioxidant capacity for the inhibition of LDL oxidation

A novel model of peroxyl radical initiated low-density lipoprotein (LDL) oxidation (LDL oxidation model for antioxidant capacity, or LOMAC) was developed to assess the free radical scavenging capacity of antioxidants and the extracts of natural products. A water-soluble free radical initiator, 2,2'-azobis(amidinopropane) dihydrochloride, was used at physiological temperature (37 degrees C) to generate peroxyl radicals to catalyze lipid oxidation of LDL isolated from human plasma samples. Headspace hexanal, a major decomposition product of LDL oxidation, was measured by a headspace gas chromatograph as an indicator of antioxidant capacity of different concentrations of pure antioxidants (vitamins C and E) and the extracts of natural products (fresh apple phytochemical extracts). All vitamin C and E and apple extract concentrations tested resulted in increasing partial suppression and delay of LDL oxidation. On the basis of the median effective dose (EC(50)) calculated for each compound or extract tested, the LOMAC value of 100 g of apple against LDL oxidation was equivalent to 1470 mg of vitamin E or to 402 mg of vitamin C. This study shows that the LOMAC assay can be routinely used to analyze or screen antioxidants or phytochemical extracts against LDL oxidation to prevent cardiovascular disease. The food-specific LOMAC values will be very useful as a new alternative biomarker for future epidemiological studies of cardiovascular disease.     

Novel compounds from Piper methysticum Forst (Kava Kava) roots and their effect on cyclooxygenase enzyme

Milled Piper methysticum roots were extracted sequentially with hot water and methanol. Cyclooxygenase (COX) enzyme inhibitory assay directed purification of the methanol extract yielded bornyl esters of 3,4-methylenedioxy cinnamic acid (1) and cinnamic acid (2), pinostrobin (3), flavokawain B (4), and 5,7-dimethoxyflavanone (5). The structures of compounds 1-5 were accomplished by spectral experiments. The aqueous extract contained previously reported kava lactones, as confirmed by TLC analysis. Compounds 3 and 5 were isolated for the first time from kava kava roots. Compound 4 showed the highest COX-I inhibitory activity at 100 microg/mL. All the compounds tested gave good COX-I and moderate COX-II enzyme inhibitory activities at 100 microg/mL. This is the first report of COX-I and -II inhibitory activities for compounds 1-5.     

Inhibitory effect of hot-water extract from dried fruit of Crataegus pinnatifida on low-density lipoprotein (LDL) oxidation in cell and cell-free systems

The dried fruit of Crataegus pinnatifida, a local soft drink material and medical herb, was found to possess potential against oxidative stress. In the preliminary study, the antioxidant potential of a hot-water extract obtained from the dried fruit of C. pinnatifida (CF-H) was evaluated in terms of its capacity of quenching 1,1-diphenyl-2-picrylhydrazyl free radicals (EC(50) = 0.118 mg/mL). After content analysis, it was found that CF-H is mainly composed of polyphenols including flavonoids (6.9%), procyanidins (2.2%), (+)-catechin (0.5%), and (-)-epicatechin (0.2%). The antioxidative bioactivity of CF-H had been assess previously using the models of CuSO(4) as cell-free system and sodium nitroprusside (SNP) plus macrophage RAW 264.7 cells as cell system to induce human low-density lipoprotein oxidation. CF-H was found to inhibit relative electrophoretic mobility and thiobarbituric acid reactive substances at the concentration of 0.5-1.0 mg/mL in the cell-free system and at 0.01-0.10 mg/mL in the cell system. Furthermore, it was found that CF-H decreased the SNP-induced cell lipid peroxidation and reduced glutathione depletion.     

Phenolic compounds from the leaf extract of artichoke (Cynara scolymus L.) and their antimicrobial activities

A preliminary antimicrobial disk assay of chloroform, ethyl acetate, and n-butanol extracts of artichoke (Cynara scolymus L.) leaf extracts showed that the n-butanol fraction exhibited the most significant antimicrobial activities against seven bacteria species, four yeasts, and four molds. Eight phenolic compounds were isolated from the n-butanol soluble fraction of artichoke leaf extracts. On the basis of high-performance liquid chromatography/electrospray ionization mass spectrometry, tandem mass spectrometry, and nuclear magnetic resonance techniques, the structures of the isolated compounds were determined as the four caffeoylquinic acid derivatives, chlorogenic acid (1), cynarin (2), 3,5-di-O-caffeoylquinic acid (3), and 4,5-di-O-caffeoylquinic acid (4), and the four flavonoids, luteolin-7-rutinoside (5), cynaroside (6), apigenin-7-rutinoside (7), and apigenin-7-O-beta-D-glucopyranoside (8), respectively. The isolated compounds were examined for their antimicrobial activities on the above microorganisms, indicating that all eight phenolic compounds showed activity against most of the tested organisms. Among them, chlorogenic acid, cynarin, luteolin-7-rutinoside, and cynaroside exhibited a relatively higher activity than other compounds; in addition, they were more effective against fungi than bacteria. The minimum inhibitory concentrations of these compounds were between 50 and 200 microg/mL.     

Antioxidative and antiproliferative properties of selected barley (Hordeum vulgarae L.) cultivars and their potential for inhibition of low-density lipoprotein (LDL) cholesterol oxidation

Aqueous methanolic extracts of whole kernels from six different barley cultivars, namely, Falcon, AC Metcalfe, Tyto, Tercel, Phoenix, and Peregrine, were examined for their total phenolic content (TPC), oxygen radical scavenging capacity (ORACFL), hydroxyl radical scavenging capacity (HORACFL), potency in prevention of lipid oxidation using the Rancimat method, efficacy in inhibition of Cu(II)-induced human LDL cholesterol oxidation, and antiproliferative activities using Caco-2 colorectal adenocarcinoma cell line. Total phenolic content as measured by Folin-Ciocalteu's method ranged from 0.68 to 1.19 mg of ferulic acid equiv/g of defatted material, whereas ORACFL and HORACFL values were 11.28-19.10 and 9.06-12.99 micromol of Trolox equiv/g of defatted material, respectively. Protection factor (PF), a measure of the effect of extracts on the prevention of oxidation of stripped corn oil as measured by Rancimat, ranged from 0.97 to 1.59. Furthermore, barley extracts showed 19.64-33.93% inhibition against Cu(II)-induced human LDL cholesterol oxidation at a final concentration of 0.02 mg/mL. The proliferation of Caco-2 colon cancer cells was significantly (p < 0.05) inhibited in a dose-dependent fashion in the presence of all barley extracts tested at the end of the day 4 of incubation. At the end of day 4, barley extracts rendered 29.3-51.2 and 9.3-15.9% inhibition of cell proliferation at 0.5 and 0.05 mg/mL, respectively. Phenolic extracts from whole barley kernel tested possessed high antioxidant, antiradical, and antiproliferative potentials. Therefore, inclusion of whole barley into the daily diet may render beneficial health benefits.     

Antioxidative activity of sulfur-containing compounds in Allium species for human low-density lipoprotein (LDL) oxidation in vitro

Sulfur-containing compounds contributing to health promotion in Allium species are produced via enzymic and thermochemical reactions. Sulfur-containing amino acids and volatile organosulfur compounds were prepared for an antioxidative assay. The inhibitory activity of S-alk(en)yl-l-cysteines and their sulfoxides, volatile alk(en)yl disulfides and trisulfides, and vinyldithiins in Allium species against lipid hydroperoxide (LOOH) formation in human low-density lipoprotein (LDL) was examined. It was elucidated that the alk(en)yl substituents (methyl, propyl, and allyl) and the number of sulfur atoms in the compounds were important for the antioxidative activity. 3,4-Dihydro-3-vinyl-1,2-dithiin, which is produced by a thermochemical reaction of allyl 2-propenethiosulfinate, exhibited the highest antioxidative activity of human LDL among sulfur-containing compounds.     

Unsaturated fatty alcohol derivatives of olive oil phenolic compounds with potential low-density lipoprotein (LDL) antioxidant and antiobesity properties

A new route for the synthesis of fatty alcohol derivatives of hydroxytyrosol and other olive oil phenolic compounds was developed to allow the preparation of unsaturated derivatives. The biological activity of synthesized compounds was evaluated. Most of the compounds presented a significant antioxidant activity on low-density lipoprotein (LDL) particles. The activity of the tested products was significantly influenced by the number and position of unsaturations as well as modifications on the polar head of the synthesized compounds. Some of them presented modulation of food intake in rats and, due to their molecular similarity with CB(1) endogenous ligands, the endocannabinoid system and PPAR-α were also evaluated as potential targets. The pharmacodynamics could not be totally explained by CB(1) and PPAR-α receptor interactions because only two of the four compounds with biological activity showed a CB(1) activity and all of them presented low PPAR-α affinity, not justifying its whole in vivo activity. The hydroxytyrosol linoleylether (7) increased LDL resistance to oxidation with a capacity similar to that of hydroxytyrosol and was the most active in vivo compound with a hypophagic effect comparable to that of oleoylethanolamine. We consider that this compound could be a good lead compound for future drug development in obesity treatments.     

Cyclooxygenase inhibitory and antioxidant compounds from the mycelia of the edible mushroom Grifola frondosa

The bioassay-guided isolation and purification of the hexane extract of the cultured mycelia of Grifola frondosa led to the characterization of a fatty acid fraction and three compounds, ergosterol (1), ergostra-4,6,8(14),22-tetraen-3-one (2), and 1-oleoyl-2-linoleoyl-3-palmitoylglycerol (3). The composition of fatty acid fraction was confirmed as palmitic, oleic, and linoleic acids by GC-MS and by comparison with the retention values of authentic samples. The structures of compounds 1-3 were established by spectroscopic methods. The fatty acid fraction and compounds 1-3 showed cyclooxygenase (COX) enzyme inhibitory and antioxidant activities. The inhibition of COX-1 enzyme by the fatty acid fraction and compounds 1-3 at 250 microg/mL were 98, 37, 55, and 67%, respectively. Similarly, COX-2 enzyme activity was reduced by fatty acid fraction and compounds 1-3 at 250 microg/mL by 99, 37, 70, and 4%, respectively. The inhibitions of liposome peroxidation by the fatty acid fraction and compounds 1 and 2 at 100 microg/mL were 79, 48, and 42%, respectively. This is the first report of compounds 2 and 3 from the cultured mycelia of G. frondosa. The COX inhibitory activities of compounds 1-3 are reported here for the first time.     

Inhibition of low-density lipoprotein oxidation and oxidative burst in polymorphonuclear neutrophils by caffeic acid and hispidin derivatives isolated from sword brake fern (Pteris ensiformis Burm.)

Several antioxidant compounds have been previously identified from sword brake fern (Pteris ensiformis Burm.) by DPPH bleaching and Trolox equivalent antioxidant capacity (TEAC) analyses. Among the isolates, 7-O-caffeoylhydroxymaltol 3-O-beta-D-glucopyranoside and hispidin 4-O-beta- D-glucopyranoside [6-(3,4-dihydroxystyryl)-4-O-beta-D-glucopyranoside-2-pyrone] were two new compounds. The aim of this study is to elucidate the possible effect of the aqueous extract of sword brake fern (SBF) and these two compounds in preventing atherosclerosis. The results demonstrated that SBF and these two compounds strongly inhibited Cu2+-mediated low-density lipoprotein (LDL) oxidation measured by thiobarbituric acid-reactive substances assay (TBARS), conjugated diene production, and relative electrophoretic mobility. The commercial antioxidant dl-alpha-tocopherol showed lower antioxidant activity than these two compounds at the same molecular concentration. SBF and these two compounds also suppressed N-formylmethionyl-leucylphenylalanine (fMLP)-stimulated reactive oxygen species (ROS) production in human polymorphonuclear neutrophils (PMN). These findings indicate that sword brake fern may prevent atherosclerosis via inhibition of both LDL oxidation and ROS production.     

Antioxidant capacity of oat (Avena sativa L.) extracts. 1. Inhibition of low-density lipoprotein oxidation and oxygen radical absorbance capacity

Milled oat groat pearlings, trichomes, flour, and bran were extracted with methanol and the fractions tested in vitro for antioxidant capacity against low-density lipoprotein (LDL) oxidation and R-phycoerythrin protein oxidation in the oxygen radical absorbance capacity (ORAC) assay. The oxidative reactions were generated by 2,2'-azobis(2-amidinopropane) HCl (AAPH) or Cu(2+) in the LDL assay and by AAPH or Cu(2+) + H(2)O(2) in the ORAC assay and calibrated against a Trolox standard to calculate Trolox equivalents (1 Trolox equivalent = 1 TE = activity of 1 micromol of Trolox). The antioxidant capacity of the oat fractions was generally consistent with a potency rank of pearlings (2.89-8.58 TE/g) > flour (1.00-3.54 TE/g) > trichome (1.74 TE/g) = bran (1.02-1.62 TE/g) in both LDL and ORAC assays regardless of the free radical generator employed. A portion of the oat antioxidant constituents may be heat labile as the greatest activity was found among non-steam-treated pearlings. The contribution of oat tocols from the fractions accounted for <5% of the measured antioxidant capacity. AAPH-initiated oxidation of LDL was inhibited by the oat fractions in a dose-dependent manner, although complete suppression was not achieved with the highest doses tested. In contrast, Cu(2+)-initiated oxidation of LDL stimulated peroxide formation with low oat concentrations but completely inhibited oxidation with higher doses. Thus, oats possess antioxidant capacity most of which is likely derived from polar phenolic compounds in the aleurone.     

Protective effects of berberine against low-density lipoprotein (LDL) oxidation and oxidized LDL-induced cytotoxicity on endothelial cells

The oxidative modification of low-density lipoprotein (LDL) is thought to have a central role in the pathogenesis of atherogenesis. Berberine, a natural constituent of plants of the genera Coptis and Berberis, has several anti-inflammation and anticancer biological effects. However, its protective effects on LDL oxidation and endothelial injury induced by oxLDL remain unclear. In this study, we evaluated the antioxidative activity of berberine and how berberine rescues human umbilical vein endothelial cells (HUVECs) from oxidized LDL (oxLDL)-mediated dysfunction. The antioxidative activity of berberine was defined by the relative electrophoretic mobility of oxLDL, fragmentation of ApoB, and malondialdehyde production via the Cu(2+)-mediated oxidation of LDL. Berberine also inhibited the generation of ROS and the subsequent mitochondrial membrane potential collapse, chromosome condensation, cytochrome C release, and caspase-3 activation induced by oxLDL in HUVECs. Our results suggest that berberine may protect LDL oxidation and prevent oxLDL-induced cellular dysfunction.     

Topoisomerase I and II enzyme inhibitory aqueous extract of Ardisia compressa and ardisin protect against benomyl oxidation of hepatocytes

Tea preparations of Ardisia compressa (AC) have been used in folk medicine against liver disorders. The objective of this study was to evaluate the in vitro topoisomerase I and II enzyme inhibition and the antioxidant effect of an aqueous extract from dry leaves of AC and a pure component (ardisin) purified from AC on benomyl (Be)-induced cytotoxicity in primary culture rat hepatocytes. Lipid peroxidation (malondialdehyde), antioxidant enzyme activities of glutathione reductase, glutathione peroxidase, and superoxide dismutase, and glutathione levels were studied. Topoisomerase I and II enzyme inhibition was used to guide purification of ardisin, which was purified using TLC, MPLC, and preparative and analytical HPLC methods. Benomyl increased malondialdehyde (58% change in comparison to the control) and glutathione peroxidase (10%), producing a significant consumption of endogenous antioxidant glutathione (65%, P < 0.05). A 94% hepatocyte protection was observed when cells were first exposed to ardisin (0.27 microg/mL), followed by Be (35 microg/mL). Cell protection by the tea extract of AC (AE) was greater than that by (-)-epigallocatechin 3-gallate (EGCG). Ardisin showed a clear inhibition of topoisomerases I and II catalytic activity in Saccharomyces cerevisiae mutant cells JN 394, JN394t(-)(1), and JN394t-(2)(-)(5). The potency of ardisin was superior to that of AE and EGCG as an antioxidant, protecting rat hepatocytes when exposed to Be. On the basis of the effective concentrations of equivalents to [+]catechin found in the present study, it can be estimated that, in order to gain antioxidative protection, a person would need to ingest approximately 1 L of AC tea per day, with a total content of 10.8 g of plant material.     

Bioguided isolation and identification of the nonvolatile antioxidant compounds from fennel (Foeniculum vulgare Mill.) waste

A bioguided isolation of an aqueous extract of fennel waste led to the isolation of 12 major phenolic compounds. Liquid chromatography coupled to atmospheric pressure chemical ionization mass spectrometry (LC/UV/APCI-MS) combined with spectroscopic methods (NMR) was used for compound identification. Radical scavenging activity was tested using three methods: DPPH*, superoxide nitro-blue tetrazolium hypoxanthine/xanthine oxidase, and *OH/luminol chemiluminescence. In addition to products described in the literature, eight antioxidant compounds were isolated and identified for the first time in fennel: 3-caffeoylquinic acid, 4-caffeoylquinic acid, 1,5-O-dicaffeoylquinic acid, rosmarinic acid, eriodictyol-7-O-rutinoside, quercetin-3-O-galactoside, kaempferol-3-O-rutinoside, and kaempferol-3-O-glucoside. The structures of eriodictyol-7-O-rutinoside and quercetin-3-O-glucuronide were completely elucidated by two-dimensional NMR experiments. The isolated compounds exhibited a strong antiradical scavenging activity, which may contribute to the interpretation of the pharmacological effects of fennel.     

Milk thistle extracts inhibit the oxidation of low-density lipoprotein (LDL) and subsequent scavenger receptor-dependent monocyte adhesion

Silymarin encompasses a group of flavonolignans that are extracted from Silybum marianum (Asteraceae) fruits. The silymarins have previously been reported to lower low-density lipoprotein (LDL) levels associated with high-fat diets. The present study reports the efficacy of the silymarins in inhibiting oxidized low-density lipoprotein (oxLDL) generation and subsequent scavenger receptor (SR) mediated monocyte adherence to oxLDL. The flavonolignans that comprise silymarin include silichristin (SC), silidianin (SD), silibinin (SBN), and isosilibinin (IS). These flavonolignans (300 microM) lowered oxLDL generation, measured by the thiobarbituric acid-reacting substances (TBARS) assay, by 60.0, 28.1, 60.0, and 30.1%, respectively. SBN treatment of LDL in the presence of copper sulfate (CuSO 4) resulted in a significant dose-dependent inhibition of monocyte adhesion. Inhibition was paralleled by a decrease in binding of anti-oxLDL antibodies recognized by U937 monocyte Fc gamma receptors (FcgammaR). These results showed that silymarin and SBN, likely through antioxidant and free radical scavenging mechanisms of action, inhibit the generation of oxLDL and oxidation-specific neoepitopes recognized by SR and FcgammaR expressed on monocytes/macrophages.     

Fermentation with Aspergillus awamori enhanced contents of amino nitrogen and total phenolics as well as the low-density lipoprotein oxidation inhibitory activity of black soybeans

A solid fermentation was performed on black soybeans with Aspergillus awamori. The effects of fermentation on the contents of total phenolics and amino nitrogen and on the inhibitory effect on low-density lipoprotein (LDL) oxidation of black soybeans were examined. Results revealed that fermentation significantly enhanced the LDL oxidation inhibitory activity and total phenolics and amino nitrogen contents of black soybeans. The increased content of amino nitrogen was closely related to the enhanced LDL oxidation inhibitory activity of fermented black soybeans and its water extract. Fermentation temperature and length affected the LDL oxidation inhibitory effect exerted by the prepared fermented black soybeans. The A. awamori-fermented black soybean prepared at 30 °C for 3 days exhibited the highest inhibitory effect on LDL oxidation. The bioactive principles related to the inhibitory effect on LDL oxidation in black soybeans, regardless of fermentation, could be most efficiently extracted with water rather than 80% methanol or 80% ethanol.     

黄瓜地上部水浸液对番茄的化感抑制效应

采用培养试验研究了黄瓜地上部水浸液对番茄的化感作用及其生理生化机理,并用试管法对其水浸液的主要化学成分进行定性分析.结果表明,黄瓜地上部水浸液含有生物碱、酚类、有机酸、皂甙、甾体、多糖、蛋白质和油脂,不含鞣质.黄瓜地上部水浸液处理显著抑制番茄种子萌发过程中的呼吸速率和α-淀粉酶活性,使种子萌发率和萌发指数降低,其抑制作用随处理浓度提高而增强.在番茄幼苗生长过程中浇灌黄瓜地上部水浸液提高了番茄叶片中超氧阴离子(O2)、H2O2和丙二醛(MDA)的含量及质膜透性;降低了过氧化氢酶(CAT)和过氧化物酶(POD)活性,还原型谷胱甘肽(GSH)和抗坏血酸(ASA)含量提高,而超氧化物歧化酶(SOD)和抗坏血酸过氧化物酶(APX)活性随处理浓度增加先升后降;叶绿素含量、净光合速率(Pn)、气孔导度(Gs)和蒸腾速率(Tr)降低,胞间CO2浓度(Ci升高,幼苗干重下降.叶绿素荧光动力学资料显示,番茄叶片的初始荧光(Fo)随着处理浓度提高而增大,最大荧光(Fm)、PSⅡ潜在光化学效率(Fv/Fo)、PSⅡ原初光能转换效率(Fv/Fm)、PSⅡ光合电子传递量子效率(ΦPSⅡ)、光化学猝灭系数(qP)、表观光合电子传递速率(ETR)和光化学速率(PCR)下降,非光化学猝灭系数(NPQ)先升后降,表明叶片光合机构受到伤害.     

Serotonin derivatives, major safflower (Carthamus tinctorius L.) seed antioxidants, inhibit low-density lipoprotein (LDL) oxidation and atherosclerosis in apolipoprotein E-deficient mice

The effects of defatted safflower seed extract and its phenolic constituents, serotonin derivatives, on atherosclerosis were studied. Ethanol-ethyl acetate extract of safflower seeds (SSE) inhibited low-density lipoprotein (LDL) oxidation induced in vitro by an azo-containing free-radical initiator V70 or copper ions. Two serotonin derivatives [N-(p-coumaroyl)serotonin, CS; N-feruloylserotonin, FS] and their glucosides were identified as the major phenolic constituents of the extract. The study with chemically synthesized materials revealed that a majority of the antioxidative activity of SSE was attributable to the aglycones of these two serotonin derivatives. Orally administered CS and FS suppressed CuSO(4)-induced plasma oxidation ex vivo. Long-term (15 week) dietary supplementation of SSE (1.0 wt %/wt) and synthetic serotonin derivatives (0.2-0.4%) significantly reduced the atherosclerotic lesion area in the aortic sinus of apolipoprotein E-deficient mice (29.2-79.7% reduction). The plasma level of both lipid peroxides and anti-oxidized LDL autoantibody titers decreased concomitantly with the reduction of lesion formation. Serotonin derivatives were detected as both intact and conjugated metabolites in the plasma of C57BL/6J mice fed on 1.0% SSE diet. These findings demonstrate that serotonin derivatives of SSE are absorbed into circulation and attenuate atherosclerotic lesion development possibly because of the inhibition of oxidized LDL formation through their strong antioxidative activity.     

Enhancement of Rooibos (Aspalathus linearis) aqueous extract and antioxidant yield with fungal enzymes

The leaves and stems of the Rooibos plant ( Aspalathus linearis) are used for the production of an herbal tea known for its health promoting properties, which have been linked to its flavonoid content but which is substantially reduced by the traditional processing method employed. Selected food-grade fungi were screened for their potential to improve the yield of soluble matter extracted from rooibos plant material. Fungal cocktails of hydrolyzing enzymes enhanced either the yield of soluble solids ( Lentinula edodes and Rhizopus oryzae cultured in yeast peptone-wheat straw medium) or the yield in antioxidants from fermented rooibos ( R. oryzae cultured in potato dextrose or yeast peptone-wheat straw medium). When applied to green rooibos, L. edodes (cultured in yeast peptone-wheat straw medium) enhanced the release of soluble solids as well as color formation, leading to semifermented rooibos with a relatively high aspalathin content, compared to fermented rooibos.