Propyl gallate induces human pulmonary fibroblast cell death through the regulation of Bax and caspase-3.

Propyl gallate (PG) has been found to exert an inhibitory effect on the growth of different cell types, including lung cancer cells. However, little is known about the cytotoxicological effects of PG specifically on normal primary lung cells. The current study examined the cellular effects and cell death resulting from PG treatment in human pulmonary fibroblast (HPF) cells. DNA flow cytometry results demonstrated that PG (100-1,600 µM) had a significant impact on the cell cycle, leading to G1 phase arrest. Notably, 1,600 µM PG slightly increased the number of sub-G1 cells. Additionally, PG (400-1,600 µM) resulted in the initiation of cell death, a process that coincided with a loss of mitochondrial membrane potential (MMP; ΔΨm). This loss of MMP (ΔΨm) was evaluated using a FACS cytometer. In PG-treated HPF cells, inhibitors targeting pan-caspase, caspase-3, caspase-8, and caspase-9 showed no significant impact on the quantity of annexin V-positive and MMP (ΔΨm) loss cells. The administration of siRNA targeting Bax or caspase-3 demonstrated a significant attenuation of PG-induced cell death in HPF cells. However, the use of siRNAs targeting p53, Bcl-2, or caspase-8 did not exhibit any notable effect on cell death. Furthermore, none of the tested MAPK inhibitors, including MEK, c-Jun N-terminal kinase (JNK), and p38, showed any impact on PG-induced cell death or the loss of MMP (ΔΨm) in HPF cells. In conclusion, PG induces G1 phase arrest of the cell cycle and cell death in HPF cells through apoptosis and/or necrosis. The observed HPF cell death is mediated by the modulation of Bax and caspase-3. These findings offer insights into the cytotoxic and molecular effects of PG on normal HPF cells.

Transcriptome analysis reveals the effect of propyl gallate on kiwifruit callus formation.

KEY MESSAGE: Exploring the potential action mechanisms of reactive oxygen species during the callus inducing, they can activate specific metabolic pathways in explants to regulate callus development. Reactive oxygen species (ROS) play an important role in the regulation of plant growth and development, but the mechanism of their action on plant callus formation remains to be elucidated. To address this question, kiwifruit was selected as the explant for callus induction, and the influence of ROS on callus formation was investigated by introducing propyl gallate (PG) as an antioxidant into the medium used for inducing callus. The results have unveiled that the inclusion of PG in the medium has disturbed the equilibrium of ROS during the formation of the kiwifruit callus. We selected the callus that was induced by the addition of 0.05 mmol/L PG to the MS medium. The callus exhibited a significant difference in the amount compared to the control medium without PG. The callus induced by the MS medium without PG was used as the control for comparison. KEGG enrichment indicated that PG exposure resulted in significant differences in gene expression in related pathways, such as phytohormone signaling and glutathione in kiwifruit callus. Weighted gene co-expression analysis indicated that the pertinent regulatory networks of both ROS and phytohormone signaling were critical for the establishment of callus in kiwifruit leaves. In addition, during the process of callus establishment, the ROS level of the explants was also closely related to the genes for transmembrane transport of substances, cell wall formation, and plant organ establishment. This investigation expands the theory of ROS-regulated callus formation and presents a new concept for the expeditious propagation of callus in kiwifruit.

High-dose synthetic phenolic antioxidant propyl gallate impairs mouse oocyte meiotic maturation through inducing mitochondrial dysfunction and DNA damage.

Propyl gallate (PG) is one of the most widely used antioxidants in food products, cosmetics and pharmaceutical industries. Increased research has suggested that exposure to PG influences reproductive health in humans and animals. However, until now, it has not yet been confirmed whether PG would impact oocyte quality. In this study, the hazardous effects of PG on oocyte meiotic maturation were investigated in mice. The findings showed that PG exposure compromises oocyte meiosis by inducing mitochondrial stress which activates apoptosis to trigger oocyte demise. Moreover, DNA damage was significantly induced in PG-treated oocytes, which might be another cause of oocyte developmental arrest and degeneration. Besides, the level of histone methylation (H3K27me2 and H3K27me3) in oocyte was also significantly increased by PG exposure. Furthermore, PG-induced oxidative stress was validated by the increased level of reactive oxygen species (ROS), which might be the underlying reason for these abnormities. In conclusion, the foregoing findings suggested that PG exposure impaired oocyte meiotic maturation by yielding mitochondrial stress to activate apoptosis, inducing DNA damage and oxidative stress, and altering histone methylation level.

Solvent-free enzymatic synthesis of 1,2-dipalmitoylgalloylglycerol: Characterization and optimization of reaction condition.

A novel diacylglycerol-based galloyl structured lipid, 1,2-dipalmitoylgalloylglycerol (DPGG), was synthesized using the enzymatic transesterification of propyl gallate (PG) and tripalmitin under solvent-free condition. An immobilized and commercially available food-grade Candida antarctica lipase B, Lipozyme® 435, was used as the biocatalyst. The reaction variables that affect the yield of DPGG were optimized using a 33 full factorial design. At 70 °C, DPGG was obtained at a yield of 33.0 ± 2.0% with PG conversion at 44.8 ± 1.8% when the following condition was used: 25 substrate molar ratio of tripalmitin to PG, 120 h reaction time, and 25% enzyme load relative to the total substrate weight. The structure of reaction product was elucidated using Fourier-transform infrared spectroscopy (FT-IR), electrospray ionization high-resolution accurate-mass tandem mass spectrometry (ESI-HRAM-MS/MS), and 1D and 2D nuclear magnetic resonance spectroscopy (NMR). The effects of different lipases and galloyl donors/acceptors on the transesterification were also investigated.

Extracellular Tannase from Aspergillus ochraceus: Influence of the Culture Conditions on Biofilm Formation, Enzyme Production, and Application.

Aspergillus ochraceus biofilm, developed on an inert support, can produce tannase in Khanna medium containing 1.5% (w/v) tannic acid as the carbon source, at an initial pH of 5.0, for 72 h at 28 °C. Addition of 0.1% yeast extract increased enzyme production. The enzyme in the crude filtrate exhibited the highest activity at 30 °C and pH 6.0. At 50 °C, the half-life was 60 min and 260 min at pH 6.0. In general, addition of detergents and surfactants did not affect tannase activity significantly. Tannase has potential applications in various biotechnological processes such as the production of propyl gallate and in the treatment of tannin-rich effluents. The content of tannins and total phenolic compounds in effluents from leather treatment was reduced by 56-83% and 47-64%, respectively, after 2 h of enzyme treatment. The content of tannins and total phenolic compounds in the sorghum flour treated for 120 h with tannase were reduced by 61% and 17%, respectively. Interestingly, the same A. ochraceus biofilm was able to produce tannase for three sequential fermentative process. In conclusion, fungal biofilm is an interesting alternative to produce high levels of tannase with biotechnological potential to be applied in different industrial sectors.

Effect of the structure of gallic acid and its derivatives on their interaction with plant ferritin.

Gallic acid and its derivatives co-exist with protein components in foodstuffs, but there is few report on their interaction with proteins. On the other hand, plant ferritin represents not only a novel class of iron supplement, but also a new nanocarrier for encapsulation of bioactive nutrients. However, plant ferritin is easy to be degraded by pepsin in the stomach, thereby limiting its application. Herein, we investigated the interaction of gallic acid and its derivatives with recombinant soybean seed H-2 ferritin (rH-2). We found that these phenolic acids interacted with rH-2 in a structure-dependent manner; namely, gallic acid (GA), methyl gallate (MEGA) and propyl gallate (PG) having three HO groups can bind to rH-2, while their analogues with two HO groups cannot. Consequently, such binding largely inhibited ferritin degradation by pepsin. These findings advance our understanding of the relationship between the structure and function of phenolic acids.

Kinetic studies of bovine serum albumin interaction with PG and TBHQ using surface plasmon resonance.

Propyl gallate (PG) and tert-butylhydroquinone (TBHQ) are examples of phenolic antioxidant agents, which have widespread uses in food industry. In this study, for the first time, we report on the interaction of PG and TBHQ with bovine serum albumin (BSA) using surface plasmon resonance (SPR). In order to modify Au slide with carboxyl functional group, 11-mercaptoundecanoic acid (MUA) was used. After activation of carboxylic groups, BSA was immobilized onto the MUA through both covalent amide bond and electrostatic binding formation. The SPR analysis showed dose-response sensograms of BSA upon increasing concentration of PG and TBHQ. At pH 4.5, the equilibrium dissociation constant or affinity unit (KD) for PG and TBHQ were 1.89e(-10) and 1.49e(-10) and at pH 7.5 were 4.74e(-10) and 1.83e(-9), respectively. The smaller amount of KD demonstrated high food additive molecules affinity to BSA. Based on these findings, it can be concluded that PG and TBHQ molecules can interact with BSA and effectively distributed within the body. Besides, SPR can be considered as useful automatic tool for quantification of PG and TBHQ interaction with serum albumin and it can deliver precise real-time kinetic data.

Enzymatic Glycosylation of Phenolic Antioxidants: Phosphorylase-Mediated Synthesis and Characterization.

Although numerous biologically active molecules exist as glycosides in nature, information on the activity, stability, and solubility of glycosylated antioxidants is rather limited to date. In this work, a wide variety of antioxidants were glycosylated using different phosphorylase enzymes. The resulting antioxidant library, containing α/ß-glucosides, different regioisomers, cellobiosides, and cellotriosides, was then characterized. Glycosylation was found to significantly increase the solubility and stability of all evaluated compounds. Despite decreased radical-scavenging abilities, most glycosides were identified to be potent antioxidants, outperforming the commonly used 2,6-bis(1,1-dimethylethyl)-4-methylphenol (BHT). Moreover, the point of attachment, the anomeric configuration, and the glycosidic chain length were found to influence the properties of these phenolic glycosides.

Gallic acid-based alkyl esters synthesis in a water-free system by celite-bound lipase of Bacillus licheniformis SCD11501.

Gallic acid (3, 4, 5- trihydroxybenzoic acid) is an important antioxidant, anti-inflammatory, and radical scavenging agent. In the present study, a purified thermo-tolerant extra-cellular lipase of Bacillus licheniformis SCD11501 was successfully immobilized by adsorption on Celite 545 gel matrix followed by treatment with a cross-linking agent, glutaraldehyde. The celite-bound lipase treated with glutaraldehyde showed 94.8% binding/retention of enzyme activity (36 U/g; specific activity 16.8 U/g matrix; relative increase in enzyme activity 64.7%) while untreated matrix resulted in 88.1% binding/retention (28.0 U/g matrix; specific activity 8.5 U/g matrix) of lipase. The celite-bound lipase was successfully used to synthesis methyl gallate (58.2%), ethyl gallate (66.9%), n-propyl gallate (72.1%), and n-butyl gallate (63.8%) at 55(o) C in 10 h under shaking (150 g) in a water-free system by sequentially optimizing various reaction parameters. The low conversion of more polar alcohols such as methanol and ethanol into their respective gallate esters might be due to the ability of these alcohols to severely remove water from the protein hydration shell, leading to enzyme inactivation. Molecular sieves added to the reaction mixture resulted in enhanced yield of the alkyl ester(s). The characterization of synthesised esters was done through fourier transform infrared (FTIR) spectroscopy and (1) H NMR spectrum analysis.

Metabolomic analysis reveals mechanism of antioxidant butylated hydroxyanisole on lipid accumulation in Crypthecodinium cohnii.

The heterotrophic dinoflagellate alga Crypthecodinium cohnii is known to accumulate lipids with a high fraction of docosahexaenoic acid (DHA). In this study, we first evaluated two antioxidant compounds, butylated hydroxyanisole (BHA) and propyl gallate (PG), for their effects on lipid accumulation in C. cohnii. The results showed that antioxidant BHA could increase lipid accumulation in C. cohnii by 8.80% at a final concentration of 30 µM, while PG had no obvious effect on lipid accumulation at the tested concentrations. To decipher the molecular mechanism responsible for the increased lipid accumulation by BHA, we employed an integrated GC-MS and LC-MS metabolomic approach to determine the time-series metabolic profiles with or without BHA, and then subjected the metabolomic data to a principal component analysis (PCA) and a weighted gene coexpression network analysis (WGCNA) network analyses to identify the key metabolic modules and metabolites possibly relevant to the increased lipid accumulation. LC-MS analysis showed that several metabolites, including NADPH, could be important for the stimulation role of BHA on lipid accumulation. Meanwhile GC-MS and network analyses allowed identification of eight metabolic modules and nine hub metabolites possibly relevant to the stimulation role of BHA in C. cohnii. The study provided a metabolomics view of the BHA mode of action on lipid accumulation in C. cohnii, and the information could be valuable for a better understanding of antioxidant effects on lipid accumulation in other microalgae as well.

Effects of a dietary antioxidant blend and vitamin E on fatty acid profile, liver function, and inflammatory response in broiler chickens fed a diet high in oxidants.

The aim of the current study was to determine the effects of a dietary antioxidant blend and vitamin E on fatty acid profile, inflammatory response, and liver function. Cobb 500 male broilers (n = 1,200, d 0) were randomly distributed into 6 treatments with 10 replicate floor pens. Treatments included (1) a high-oxidant diet, with vitamin E at 10 IU/kg, 3% oxidized oil, 3% polyunsaturated fatty acids (PUFA) source (HO); (2) the HO diet with vitamin E at 200 IU/kg (VE); (3) the HO diet with an antioxidant blend at 135 mg/kg (AOX); (4) the HO diet with both vitamin E at 200 IU/kg and an antioxidant blend at 135 mg/kg (VE+AOX); (5) standard control (SC); and (6) a positive control, which was the SC diet with an antioxidant blend at 135 mg/kg. The concentrations of 20:4, 20:5, 22:5, 22:6, and all the n-3 fatty acids were greater in the abdominal fat of HO, VE, AOX, and VE+AOX birds than SC and positive control birds on d 21 and 42 (P < 0.001). Compared with HO treatment, AOX and VE+AOX preserved the deposition of PUFA better (P < 0.001). The HO birds had greater concentrations of aspartate aminotransferase on d 21 and 42, and γ-glutamyl transferase on d 21, whereas AOX and VE+AOX chickens had restored γ-glutamyl transferase concentration (P < 0.01). The inflammation scores of abdominal fat of AOX and VE+AOX birds were lower than the HO on d 21 (P < 0.001). Compared with SC, the VE and VE+AOX birds exhibited greater vacuole scores on d 21 and 42 (P < 0.01). The lower vacuoles score in SC was associated with a greater expression of peroxisome proliferator activated receptor -γ and -α (P < 0.05). The expression of inflammatory genes in the liver did not differ among treatments. In conclusion, the AOX and AOX+VE diets were effective in preserving PUFA in the abdominal fat, moderately improved liver function, and reduced inflammation in fat.

Effects of a dietary antioxidant blend and vitamin E on growth performance, oxidative status, and meat quality in broiler chickens fed a diet high in oxidants.

The aim of the study was to determine the effects of a dietary antioxidant blend (AB) and vitamin E on performance, oxidative status, and meat quality. Cobb 500 male broilers (n = 1,200, d 0) were randomly distributed into 6 treatments with 10 replicate pens. Treatments included 1) HO: high oxidant diet, vitamin E at 10 IU/kg, 3% oxidized soybean oil, 3% polyunsaturated fatty acid (PUFA) source; 2) VE: the HO diet with vitamin E at 200 IU/kg; 3) AOX: the HO diet with AB at 135 mg/kg; 4) VE+AOX: the HO diet with vitamin E at 200 IU/kg and AB at 135 mg/kg; 5) SC: standard control; and 6) PC: positive control, the SC diet with AB at 135 mg/kg. From d 0 through d 21, high oxidant diet treatment birds had greater BW, ADG, and ADFI than the SC birds; the AOX birds had better G:F on d 10 and 42, and from d 0 to 42 than SC birds (P < 0.05). The plasma TBA reactive substance level was lower in the AOX birds than the VE treatment birds in all phases (P < 0.05). High oxidant diet treatment birds had greater α-1-acid glycoprotein levels on d 10 than SC and PC birds (P < 0.05). The AOX, PC, and SC birds had a greater level of uric acid than the HO and VE+AOX birds on d 10. Superoxide dismutase expression in the liver was less with the HO treatment compared with the SC treatment on d 7 (P < 0.05). The vitamin E concentration in the breast muscle was greatest in the VE birds, whereas vitamin A concentration was greater in the PC birds compared with the SC birds on d 21 (P < 0.05). Compared with VE and AOX, the HO treatment had greater drip loss (P < 0.05). In conclusion, dietary addition of AOX was effective in improving growth, moderately restored the whole body antioxidant capability, and reduced drip loss.

Production of propyl gallate in nonaqueous medium using cell-associated tannase of Bacillus massiliensis: effect of various parameters and statistical optimization.

Enzymatic synthesis of propyl gallate in an organic solvent was studied using cell-associated tannase (E.C. 3.1.1.20) of Bacillus massiliensis. Lyophilized biomass showing tannase activity was used as a biocatalyst. The influence of buffer pH and strength, water activity, temperature, biocatalyst loading, gallic acid concentration, and 1-propanol concentration was studied by the one-factor-at-a-time method. Subsequently, response surface methodology was applied based on a central composite design to determine the effects of three independent variables (biocatalyst loading, gallic acid concentration, and 1-propanol concentration) and their mutual interactions. A total of 20 experiments were conducted, and a statistical model was developed, which predicted the maximum propyl gallate yield of 20.28 µg/mL in the reaction mixture comprising 40.4 mg biocatalyst, 0.4 mM gallic acid, and 6.52 % (v/v) 1-propanol in 9.5 mL benzene at 30°C. The subsequent verification experiments established the validity of the model. Under optimal conditions, 25% conversion of gallic acid to propyl gallate was achieved on a molar basis. The absence of the need for enzyme purification and subsequent immobilization steps and good conversion efficiency makes this enzyme system an interesting one. Reports on the applications of bacterial whole cell systems for synthetic reactions in organic solvents are scarce, and perhaps this is the first report on bacterial cell-associated tannase-mediated esterification in a nonaqueous medium.

Characterization of a tannase from Emericella nidulans immobilized on ionic and covalent supports for propyl gallate synthesis.

The extracellular tannase from Emericela nidulans was immobilized on different ionic and covalent supports. The derivatives obtained using DEAE-Sepharose and Q-Sepharose were thermally stable from 60 to 75 °C, with a half life (t50) >24 h at 80 °C at pH 5.0. The glyoxyl-agarose and amino-glyoxyl derivatives showed a thermal stability which was lower than that observed for ionic supports. However, when the stability to pH was considered, the derivatives obtained from covalent supports were more stable than those obtained from ionic supports. DEAE-Sepharose and Q-Sepharose derivatives as well as the free enzyme were stable in 30 and 50 % (v/v) 1-propanol. The CNBr-agarose derivative catalyzed complete tannic acid hydrolysis, whereas the Q-Sepharose derivative catalyzed the transesterification reaction to produce propyl gallate (88 % recovery), which is an important antioxidant.

Characterization of bioimprinted tannase and its kinetic and thermodynamics properties in synthesis of propyl gallate by transesterification in anhydrous medium.

Tannase has been extensively applied to synthesize gallic acid esters. Bioimprinting technique can evidently enhance transesterification-catalyzing performance of tannase. In order to promote the practical utilization of the modified tannase, a few enzymatic characteristics of the enzyme and its kinetic and thermodynamics properties in synthesis of propyl gallate by transesterification in anhydrous medium have been studied. The investigations of pH and temperature found that the imprinted tannase holds an optimum activity at pH 5.0 and 40 °C. On the other hand, the bioimprinting technique has a profound enhancing effect on the adapted tannase in substrate affinity and thermostability. The kinetic and thermodynamic analyses showed that the modified tannase has a longer half-time of 1,710 h at 40 °C; the kinetic constants, the activation energy of reversible thermal inactivation, and the activation energy of irreversible thermal inactivation, respectively, are 0.054 mM, 17.35 kJ mol(-1), and 85.54 kJ mol(-1) with tannic acid as a substrate at 40 °C; the free energy of Gibbs (ΔG) and enthalpy (ΔH) were found to be 97.1 and 82.9 kJ mol(-1) separately under the same conditions.

Propyl gallate synthesis using acidophilic tannase and simultaneous production of tannase and gallic acid by marine Aspergillus awamori BTMFW032.

Marine Aspergillus awamori BTMFW032, recently reported by us, produce acidophilic tannase as extracellular enzyme. Here, we report the application of this enzyme for synthesis of propyl gallate by direct transesterification of tannic acid and in tea cream solubilisation besides the simultaneous production of gallic acid along with tannase under submerged fermentation by this fungus. This acidophilic tannase enabled synthesis of propyl gallate by direct transesterification of tannic acid using propanol as organic reaction media under low water conditions. The identity of the product was confirmed with thin layer chromatography and Fourier transform infrared spectroscopy. It was noted that 699 U/ml of enzyme could give 60% solubilisation of tea cream within 1 h. Enzyme production medium was optimized adopting Box-Behnken design for simultaneous synthesis of tannase and gallic acid. Process variables including tannic acid, sodium chloride, ferrous sulphate, dipotassium hydrogen phosphate, incubation period and agitation were recognized as the critical factors that influenced tannase and gallic acid production. The model obtained predicted 4,824.61 U/ml of tannase and 136.206 µg/ml gallic acid after 48 h of incubation, whereas optimized medium supported 5,085 U/ml tannase and 372.6 µg/ml of gallic acid production after 36 and 84 h of incubation, respectively, with a 15-fold increase in both enzyme and gallic acid production. Results indicated scope for utilization of this acidophilic tannase for transesterification of tannic acid into propyl gallate, tea cream solubilisation and simultaneous production of gallic acid along with tannase.

Induction of morphological changes in Ustilago maydis cells by octyl gallate.

The effects of octyl gallate on Ustilago maydis yeast cells were analysed in relation to its capacity to oxidize compounds (pro-oxidant actions). All phenolic compounds tested inhibited the alternative oxidase (AOX). However, only octyl gallate induced a morphological change in yeast cells and collapsed the mitochondrial membrane potential. In contrast to octyl gallate, propyl gallate and nordihydroguaiaretic acid caused only a negligible cell change and the membrane potential was not affected. Our findings show that structurally related phenolic compounds do not necessarily exert similar actions on target cells. Preincubation of U. maydis cells with trolox inhibited the change to pseudohyphal growth produced by octyl gallate. These results suggest that in addition to the inhibitory action of octyl gallate on the AOX, this compound induces a switch from yeast to a mycelium, probably through the formation of lipid peroxides.

Gallic acid, a metabolite of the antioxidant propyl gallate, inhibits gap junctional intercellular communication via phosphorylation of connexin 43 and extracellular-signal-regulated kinase1/2 in rat liver epithelial cells.

Propyl gallate and its metabolite, gallic acid, are widely used as antioxidants in the food industry, but they have been shown to exhibit liver toxicity and enhance carcinogenesis. In the present study, we investigated the possible undesirable effects of propyl gallate and gallic acid on gap junctional intercellular communication (GJIC), inhibition of which is closely linked to carcinogenesis. Gallic acid and propyl gallate exhibited dose-dependent free-radical-scavenging activities as determined by 1,1-diphenyl-2-picrylhydrazyl- or 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)-radical-scavenging assays, and the free-radical-scavenging activity of gallic acid was stronger than that of propyl gallate. However, using WB-F344 rat liver epithelial cells, gallic acid inhibited GJIC in a dose-dependent manner, while propyl gallate had no significant effect compared with untreated controls. The gallic-acid-induced inhibition of GJIC was reversible, with a recovery of nearly 65% after 120 min. Gallic acid induced the phosphorylation of connexin 43 (Cx43) and phosphorylation of extracellular-signal-regulated kinase1/2 (ERK1/2). The gallic-acid-induced inhibition of GJIC was attenuated by treatment with mitogen-activated protein kinase kinase inhibitors (U0126 and PD098059). U0126 blocked the gallic-acid-induced phosphorylation of Cx43 and ERK1/2, indicating that the gallic-acid-induced inhibition of GJIC is mediated by phosphorylation of Cx43 via activation of ERK1/2. In addition, gallic-acid-induced inhibition of GJIC was protected by ascorbic acid and quercetin, which might represent a simple example of the different effects of natural antioxidants in carcinogenesis.

Microencapsulated mycelium-bound tannase from Aspergillus niger: an efficient catalyst for esterification of propyl gallate in organic solvents.

Microencapsulated Aspergillus niger with mycelium-bound tannase activity was employed to investigate the esterification of propyl gallate from gallic acid and propanol in organic solvents. The effects of various organic solvents (log P: -1.0 to 6.6) on the enzymatic reactions showed that benzene (log P: 2.0) was the suitable solvent, for which the conversion reached 26.8%. The optimum catalyst concentration and water concentration was found at 25 capsules in 10 mL of benzene and 0.04 g of water/capsule. The external mass transfer effect could be eliminated at stirring speeds of 180 rpm or higher. Both substrates 1-propanol and gallic acid had significant inhibition effects on the tannase activity. Maximum molar conversion (36.2%) was achieved with 9.1% (v/v) 1-propanol and 8 mM gallic acid and decreased with increasing amounts of substrates.

Hydrogen peroxide scavenging, antioxidant and anti-radical activity of some phenolic acids.

Some water-soluble phenolic acids were investigated as antioxidants, scavengers of hydrogen peroxide (H(2)O(2)) and scavengers of 1,1-diphenyl-2-picrylhydrazyl radical (DPPH(*)). The strongest antioxidant, scavenging of H(2)O(2) and DPPH(*) radical activity was exhibited by 3,4,5-trihydroxybenzoic (gallic) acid and 1,2,3-trihydroxybenzene (pyrogallol) with three hydroxyl groups bonded to the aromatic ring in an ortho position in relation to each other. Phenolic acids with two hydroxyl groups bonded to aromatic ring in the ortho position, such as 3,4-dihydroxycinnamic (caffeic), 3,4-dihydroxybenzoic (protocatechuic) and 2,3-dihydroxybenzoic (o-pyrocatechuic) acids, showed strong antioxidant and anti-radical activity; however, it was lower than that of 3,4,5-trihydroxybenzoic acid or 1,2,3-trihydroxybenzene. 3,5-Dihydroxybenzoic (alpha-resorcylic) and 2,4-dihydroxybenzoic (beta-resorcylic) acids with two hydroxyls bonded in the meta position in relation to each other showed moderate antioxidant and low DPPH(*) and hydrogen peroxide scavenging activity. Compounds with one hydroxyl group such as 3-hydroxybenzoic, 4-hydroxyphenylacetic and 2-hydroxybenzoic (salicylic) acids, exhibited the lowest anti-radical and antioxidant activity. The results obtained show that the antioxidant and anti-radical activity of phenolic acids correlated positively with the number of hydroxyl groups bonded to the aromatic ring. The model of an ortho substitution of hydroxyl groups to the aromatic ring seems to be adequate for antioxidant and H(2)O(2) or DPPH(*) scavenging activity of phenolic acids.