OBJECTIVETo identify the mutations of the tyrosinase gene (TYR) and P gene in patients with oculocutaneous albinism (OCA).

METHODSPolymerase chain reaction (PCR) and denaturing high performance liquid chromatography (DHPLC) were applied to detect the mutations in all exons of TYR gene and P gene. Then DNA sequencing and restriction endonuclease analysis were used to confirm the mutations detected by DHPLC. Novel mutations were screened in 100 unrelated persons with normal phenotypes to exclude the possibility of polymorphism.

RESULTSTwo mutations were detected in the P gene of the three patients and none in TYR gene. Heterozygous mutation of T450M in exon 13 of the P gene was detected in patient 1. Patient 2 had a heterozygous mutation of T450M in exon 13 and a heterozygous mutation of G775R in exon 23 of the P gene. Patient 3 had a heterozygous mutation of G775R as well. Restriction endonuclease analysis of the P gene exon 13 showed that the Oli I site had partly disappeared resulting from the heterozygous mutation T450M in patient 1 and patient 2, but not in 100 unrelated individuals. The heterozygous mutation T450M is a novel mutation.

CONCLUSIONGene diagnosis of OCA can be carried out effectively by combining PCR, DHPLC, DNA sequencing and restriction endonuclease analysis.

Albinism, Oculocutaneous ; genetics ; Base Sequence ; Catechol Oxidase ; genetics ; Child, Preschool ; DNA Mutational Analysis ; Exons ; genetics ; Female ; Hermanski-Pudlak Syndrome ; genetics ; Humans ; Monophenol Monooxygenase ; genetics ; Mutation ; Young Adult

This study aims to explore the resource utilization of used fungus-growing materials produced in the cultivation of Gastrodia elata. To be specific, based on the production practice, this study investigated the recycling mechanism of used fungus-growing materials of G. elata by Phallus inpudicus. To screen edible fungi with wide adaptability, this study examined the allelopathic effects of Armillaria mellea secretions on P. impudicus and 6 kinds of large edible fungi and the activities of enzymes related to degradation of the used fungus-growing materials of G. elata. The results showed that P. impudicus can effectively degrade cellulose, hemicellulose, and lignin in used fungus-growing materials of G. elata. The cellulase activity of A. mellea was significantly higher than that of P. impudicus, and the activities of lignin peroxidase, polyphenol oxidase, and xylanase of P. impudicus were significantly higher than those of A. mellea, which was the important reason why A. mellea and P. impudicus used different parts and components of the used fungus-growing materials to absorb carbon sources and develop ecological niche differences. The growth of P. impudicus was significantly inhibited on the used fungus-growing materials of G. elata. The secretions of A. mellea had allelopathic effects on P. impudicus and other edible fungi, and the allelopathic effects were related to the concentration of allelopathy substances. The screening result showed that the growth and development of L. edodes and A. auricular were not significantly affected by 30% of A. mellea liquid, indicating that they had high resistance to the allelopathy of A. mellea. The results showed that the activities of extracellular lignin peroxidase, polyphenol oxidase, and xylanase of the two edible fungi were similar to those of P. impudicus, and the cellulase activity was higher than that of P. impudicus. This experiment can be further verified by small-scale production tests.
Agaricales ; Ascomycota ; Basidiomycota ; Catechol Oxidase ; Cellulases ; Gastrodia

Dopamine (DA), as a catecholamine neurotransmitter widely distributed in the central nervous system and the peripheral tissues, has attracted a lot of attention. Especially in recent years, DA has been found to regulate the function of the immune system, and the involvement of DA in the intestinal mucosal inflammation-related diseases has become a hot research topic. The digestive tract is an important source of peripheral DA, and DA is not only produced in the enteric nervous system and gastrointestinal epithelium, but also produced by intestinal microorganisms. In addition to the synthetases of DA, the DA contents in body tissues are also affected by the two kinds of metabolic enzymes, monoamine oxidase (MAO) and catechol-O-methyltransferase (COMT). This article reviewed the sources, metabolism, and functions of DA in digestive tract, especially focusing on the distribution and function of MAO and COMT, the enzymes degrading DA.
Catechol O-Methyltransferase ; Catechol O-Methyltransferase Inhibitors ; Dopamine ; Gastrointestinal Tract ; Monoamine Oxidase ; Monoamine Oxidase Inhibitors

There is no consensus on the content, accumulation, transformation and content determination methods of phenolic acids in fresh Salvia miltiorrhiza. In order to find out the true content of phenolic acids in fresh S. miltiorrhiza, a variety of treatment me-thods were used in this study to prepare sample solution. The content changes of phenolic acids in S. miltiorrhiza samples with different dehydration rates were investigated during drying and shade drying processes. Polyphenol oxidase(PPO) of S. miltiorrhiza was extracted and purified by ammonium sulfate precipitation and dialysis to investigate the enzymatic properties. The content of rosmarinic acid, lithosperic acid and S. nolic acid B in S. miltiorrhiza was determined by UPLC. The results showed that the content of phenolic acids in fresh S. miltiorrhiza was highest when it was homogenized with 1 mol·L~(-1) HCl solution or 1 mol·L~(-1) HCl methanol solution. There was no significant difference in the content of phenolic acids in S. miltiorrhiza with different dehydration rates, indicating that there was no correlation between phenolic acid content and dehydration rate. The optimum pH of S. miltiorrhiza PPO was 7.6 and the optimum temperature was 40 ℃. With catechol as substrate, S. miltiorrhiza PPO had the enzymatic browning reaction which was in compliance with Michaelis equation, with Michaelis constant K_m of 0.12 mol·L~(-1) and V_(max) of 588.23 U·min~(-1). The inhibitory effect of citric acid, disodium ethylenediamine tetraacetate, ascorbic acid and sodium sulfite on S. miltiorrhiza PPO increased with the increase of inhibitor concentration, and sodium sulfite showed the strongest inhibitory effect. The present study proved that there were a large number of phenolic acids in fresh S. miltiorrhiza, which were the secondary metabolite of primitive accumulation during the growth of S. miltiorrhiza, rather than the induced product of postharvest drying and dehydration stress. This study has reference value and significance for the cultivation, harvest and processing of S. miltiorrhiza.
Catechol Oxidase ; Desiccation ; Hydroxybenzoates ; Plant Roots ; Salvia miltiorrhiza

OBJECTIVEThis paper aimed to study the dynamic changes of enzyme activities and active component contents in Lonicera japonica during different blossoming stages.

METHODThe enzyme activities of phenylalanine ammonia lyase (PAL), polyphenol oxidase (PPO), peroxidase (POD) and the contents of total phenol, total flavonoids, chlorogenic acid, anthocyanins in L. japonica during different blossoming stages were determined.

RESULTThe contents of total phenolics, total flavonoids, anthocyanins decreased from the Sanqing stage to Jinhua stage while the content of chlorogenic acid increased slightly in white period, and then decreased gradually. The activities of three enzymes decreased gradually from Sanqing stage, and got to a minimum value in Yinhua stage, then increased slightly until the Jinhua stage.

CONCLUSIONThe enzyme activities of PPO and POD correlated the content of phenolic substances positively before the Jinhua stage in L. japonica. In the period of maturity, the POD activity was strengthened due to the induction of respiration and became the key enzyme to control active component content during the mature stage.

Catechol Oxidase ; metabolism ; Flowers ; Lonicera ; chemistry ; enzymology ; Peroxidase ; metabolism ; Phenols ; analysis

To investigate the enzyme properties of the black sesame polyphenol oxidase (BsPPO), a synthesized Bsppo gene was cloned into the vector pMAL-c5x and expressed in E. coli. Subsequently, the MBP fusion label in the recombinant protein was removed by protease digestion after affinity purification. The synthesized Bsppo gene contained 1 752 bp which encodes 585 amino acids with a deduced molecular weight of 65.3 kDa. Transformation of the recombinant vector into E. coli BL21(DE3) resulted in soluble expression of the fusion protein MBP-BsPPO. The enzymatic properties of the recombinant BsPPO was investigated after MBP fusion tag excision followed by affinity purification. The results demonstrated that the optimal temperature and pH for BsPPO was 25°C and 4.0, respectively. BsPPO exhibited a good stability under low temperature and acidic environment. Low-intensity short-term light exposure increased the activity of BsPPO. Cu²⁺ could improve the activity of BsPPO while Zn²⁺ and Ca²⁺ showed the opposite effect. BsPPO could catalyze the oxidation of monophenols, diphenols, and triphenols, and exhibited good catalytic activity on l-tyrosine and vanillic acid. Moreover, BsPPO exhibited high catalytic activity on black sesame metabolites, including 2-methoxy cinnamic acid, indole-3-carboxylic acid and phloretin. These results may serve as a basis for further characterization of BsPPO.
Catechol Oxidase/genetics* ; Cloning, Molecular ; Escherichia coli/metabolism* ; Recombinant Proteins/genetics* ; Sesamum/genetics*

Polyphenol oxidase(PPO) is an important antioxidant enzyme in plants. It has the functions of scavenging active oxygen and synthesizing phenols, lignin, and plant protection factors, and can enhance the plant's resistance to stress and resistance to pests and diseases. Our previous research found that Salvia miltiorrhiza PPO gene can positively regulate salvianolic acid B synthesis. In order to further explore the mechanism, a pGBKT7-PPO bait vector was constructed using the cloned S. miltiorrhiza polyphenol oxidase gene(SmPPO, GenBank accession number: KF712274.1), and verified that it had no self-activation and no toxicity. The titer of S. miltiorrhiza cDNA library constructed by our laboratory was 4.75 × 107 cfu·mL~(-1), which met the requirements for library construction. Through yeast two-hybrid test, 22 proteins that could interact with SmPPO were screened. Only yeast PAL1 and TAT interacted with SmPPO through yeast co-transformation verification. Further verification was performed by bimolecular fluorescence complementary detection(BiFC). Only TAT and SmPPO interacted, so it meant that TAT and SmPPO interacted. TAT and SmPPO were truncated according to the domain, respectively. The first 126 amino acids of SmPPO and tyrosine amino transferase(TAT) were obtained to interact on the cell membrane and chloroplast. SmPPO was obtained by subcellular localization test, which was mainly loca-lized on the nucleus and cell membrane; TAT was localized on the cell membrane. Real-time quantitative PCR results showed that the SmPPO gene was mainly expressed in roots and stems; the TAT gene was expressed in roots, and the expression level in stems and flowers was low. This article lays a solid foundation for the in-depth study of the molecular mechanism of the interaction of S. miltiorrhiza SmPPO and TAT to regulate the synthesis of phenolic substances.
Catechol Oxidase ; Gene Expression Regulation, Plant ; Gene Library ; Plant Proteins ; genetics ; Plant Roots ; Salvia miltiorrhiza ; genetics

Foliar sprays of three plant resistance inducers, including chitosan (CH), potassium sorbate (PS) (C₆H₇kO₂), and potassium bicarbonates (PB) (KHCO₃), were used for resistance inducing against Erysiphe cichoracearum DC (powdery mildew) infecting okra plants. Experiments under green house and field conditions showed that, the powdery mildew disease severity was significantly reduced with all tested treatments of CH, PS, and PB in comparison with untreated control. CH at 0.5% and 0.75% (w/v) plus PS at 1.0% and 2.0% and/or PB at 2.0% or 3.0% recorded as the most effective treatments. Moreover, the highest values of vegetative studies and yield were observed with such treatments. CH and potassium salts treatments reflected many compounds of defense singles which leading to the activation power defense system in okra plant. The highest records of reduction in powdery mildew were accompanied with increasing in total phenolic, protein content and increased the activity of polyphenol oxidase, peroxidase, chitinase, and β-1,3-glucanase in okra plants. Meanwhile, single treatments of CH, PS, and PB at high concentration (0.75%, 2.0%, and/or 3.0%) caused considerable effects. Therefore, application of CH and potassium salts as natural and chemical inducers by foliar methods can be used to control of powdery mildew disease at early stages of growth and led to a maximum fruit yield in okra plants.
Abelmoschus* ; Bicarbonates ; Catechol Oxidase ; Chitinase ; Chitosan* ; Fruit ; Peroxidase ; Phenol ; Plants* ; Potassium* ; Salts* ; Sorbic Acid

Panic disorder is one of the chronic and disabling anxiety disorders. There has been evidence for either genetic heterogeneity or complex inheritance, with environmental factor interactions and multiple single genes, in panic disorder's etiology. Linkage studies have implicated several chromosomal regions, but no research has replicated evidence for major genes involved in panic disorder. Researchers have suggested several neurotransmitter systems are related to panic disorder. However, to date no candidate gene association studies have established specific loci. Recently, researchers have emphasized genome-wide association studies. Results of two genome-wide association studies on panic disorder failed to show significant associations. Evidence exists for differences regarding gender and ethnicity in panic disorder. Increasing evidence suggests genes underlying panic disorder overlap, transcending current diagnostic boundaries. In addition, an anxious temperament and anxiety-related personality traits may represent intermediate phenotypes that predispose to panic disorder. Future research should focus on broad phenotypes, defined by comorbidity or intermediate phenotypes. Genome-wide association studies in large samples, studies of gene-gene and gene-environment interactions, and pharmacogenetic studies are needed.
Catechol O-Methyltransferase/genetics ; Cholecystokinin/genetics ; Genetic Loci ; *Genome-Wide Association Study ; Humans ; Monoamine Oxidase/genetics ; Panic Disorder/*genetics

OBJECTIVETo evaluate the enzymatic and non-enzymatic antioxidants of leaf extract from Alpinia purpurata.

METHODSOne gram of fresh leaf of Alpinia purpurata was grinded in 2 mL of 50% ethanol and centrifuged at 10,000×g at 4°C for 10 min. The supernatant obtained was used within 4 h for various enzymatic antioxidants assays like superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST), ascorbate oxidase, peroxidase, polyphenol oxidase (PPO) and non-enzymatic antioxidants such as vitamin C, total reduced glutathione (TRG) and lipid peroxidation (LPO).

RESULTSThe leaf extract of Alpinia purpurata possess antioxidants like vitamin C 472.92±6.80 μg/mg protein, GST 372.11±5.70 μmol of 1-chloro 2,4 dinitrobenzene (CDNB)-reduced glutathione (GSH) conjugate formed/min/mg protein, GPx 281.69±6.43 μg of glutathione oxidized/min/mg protein, peroxidases 173.12±9.40 μmol/g tissue, TRG 75.27±3.55 μg/mg protein, SOD 58.03±2.11 U/mg protein, CAT 46.70±2.35 μmol of H2O2 consumed/min/mg protein in high amount whereas ascorbate oxidase 17.41±2.46 U/g tissue, LPO 2.71±0.14 nmol/L of malondialdehyde formed/min/mg protein and PPO 1.14±0.11 μmol/g tissue in moderate amount.

CONCLUSIONAlpinia purpurata has the potential to scavenge the free radicals and protect against oxidative stress causing diseases. In future, Alpinia purpurata may serve as a good pharmacotherapeutic agent.

Alpinia ; chemistry ; Antioxidants ; analysis ; Catechol Oxidase ; metabolism ; Enzymes ; metabolism ; Lipid Peroxidation ; Plant Extracts ; chemistry ; Plant Leaves ; chemistry