The Inhibitory Effect of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) on the Monophenolase and Diphenolase Activities of Mushroom Tyrosinase

In the present work, we investigated the effect of non-steroidal anti-inflammatory drugs (NSAIDs) on the monophenolase and diphenolase activity of mushroom tyrosinase. The results showed that diflunisal and indomethacin inhibited both monophenolase and diphenolase activity. For monophenolase activity, the lag time was extended in the presence of diflunisal. In the presence of indomethacin, the lag time did not change. IC50 values of monophenolase activity were estimated to be 0.112 mM (diflunisal) and 1.78 mM (indomethacin). Kinetic studies of monophenolase activity revealed that both diflunisal and indomethacin were non-competitive inhibitors. For diphenolase activity, IC50 values were estimated to be 0.197 mM (diflunisal) and 0.509 mM (indomethacin). Diflunisal and indomethacin were also found to be non-competitive diphenolase inhibitors

Dormancy in wheat grain (Triticum aestivum L.) : studies on grain-coat pigment formation and abscisic acid content during the development of wheat grain of six genotypes : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Botany at Massey University

Dormancy in wheat grain has been associated with red pigmentation of the grain-coats. The development from anthesis to harvest-ripeness of two-white-grained and four red-grained genotypes of varying dormancy was investigated. Grain growth was measured as changes in fresh weight and dry matter. Dehydration to harvest-ripeness (17.5% moisture) was calculated. The developmental rates of grain of the six genotypes were similar. Dormancy-breaking germination tests showed that embryo maturity was attainted at similar stages of development of four genotypes. It appeared to be somewhat delayed in two red-grained genotypes, which also had the lowest germination rates in standard germination tests. Dormancy was estimated as the percentage of grains with mature embryos, which did not germinate in the standard germination tests. Grain of all the genotypes had a period of dormancy during development. However, in white-grained genotypes it had disappeared before harvest-ripeness was attained and it lasted only a little longer in one of the red-grained genotypes. In the other three red-grained genotypes, dormancy was prolonged for at least several weeks beyond harvest-ripeness. The concentrations of flavonoid precursors were similar in grains of all six genotypes throughout their development. Assays of crude extracts of a group of enzymes (phenolases) involved in pigment synthesis did not reveal peaks of activity associated with the appearance of mature grain-coat colour. Successive extractions of the grains showed that the pigment was probably a large flavonoid polymer. The amounts of endogenous abscisic acid in developing grains was analysed by high pressure liquid chromatography. Significant quantities of the 2-trans isomer, as well as of the common 2-cis isomer (abscisic acid) were found. The amounts did not appear to be related to either dormancy or to maturation and dehydration of the grain, as had been suggested. The mechanisns prolonging dormancy beyond harvest-ripeness in wheat grain were discussed with reference to pigmentation. It was considered that dormancy of the red-grained wheats was probably due to impermeability of the grain-coat to oxygen, possibly resulting from molecular properties of the pigment. These properties were the ability to absorb oxygen, which might prevent it reaching the embryo, and the ability to complex with the large proteins of the immature testa, which might prevent their degradation during grain development. During imbibition the complexed proteins might swell to create a physical barrier to oxygen permeation

A tyrosinase, mTyr-CNK, that is functionally available as a monophenol monooxygenase

Tyrosinase efficiently catalyzes the ortho-hydroxylation of monophenols and the oxidation of diphenols without any additional cofactors. Although it is of significant interest for the biosynthesis of catechol derivatives, the rapid catechol oxidase activity and inactivation of tyrosinase have hampered its practical utilization as a monophenol monooxygenase. Here, we prepared a functional tyrosinase that exhibited a distinguished monophenolase/diphenolase activity ratio (V max mono/ V max di = 3.83) and enhanced catalytic efficiency against L-tyrosine (k cat  = 3.33 ± 0.18 s−1, K m  = 2.12 ± 0.14 mM at 20 °C and pH 6.0). This enzyme was still highly active in ice water (>80%), and its activity was well conserved below 30 °C. In vitro DOPA modification, with a remarkably high yield as a monophenol monooxygenase, was achieved by the enzyme taking advantage of these biocatalytic properties. These results demonstrate the strong potential for this enzyme’s use as a monophenol monooxygenase in biomedical and industrial applications.113Nsciescopu

An Updated Review of Tyrosinase Inhibitors

Tyrosinase is a multifunctional, glycosylated, and copper-containing oxidase, which catalyzes the first two steps in mammalian melanogenesis and is responsible for enzymatic browning reactions in damaged fruits during post-harvest handling and processing. Neither hyperpigmentation in human skin nor enzymatic browning in fruits are desirable. These phenomena have encouraged researchers to seek new potent tyrosinase inhibitors for use in foods and cosmetics. This article surveys tyrosinase inhibitors newly discovered from natural and synthetic sources. The inhibitory strength is compared with that of a standard inhibitor, kojic acid, and their inhibitory mechanisms are discussed

Study of tyrosine and dopa enantiomers as tyrosinase substrates initiating L‐ and D‐melanogenesis pathways

Tyrosinase starts melanogenesis and determines its course, catalyzing the oxidation by molecular oxygen of tyrosine to dopa, and that of dopa to dopaquinone. Then, nonenzymatic coupling reactions lead to dopachrome, which evolves toward melanin. Recently, it has been reported that d‐tyrosine acts as tyrosinase inhibitor and depigmenting agent. The action of tyrosinase on the enantiomers of tyrosine (l‐tyrosine and d‐tyrosine) and dopa (l‐dopa and d‐dopa) was studied for the first time focusing on quantitative transient phase kinetics. Post‐steady‐state transient phase studies revealed that l‐dopachrome is formed more rapidly than d‐dopachrome. This is due to the lower values of Michaelis constants for l‐enantiomers than for d‐enantiomers, although the maximum rates are equal for both enantiomers. A deeper analysis of the inter‐steady‐state transient phase of monophenols demonstrated that the enantiomer d‐tyrosine causes a longer lag period and a lower steady‐state rate, than l‐tyrosine at the same concentration. Therefore, d‐melanogenesis from d‐tyrosine occurs more slowly than does l‐melanogenesis from l‐tyrosine, which suggests the apparent inhibition of melanin biosynthesis by d‐tyrosine. As conclusion, d‐tyrosine acts as a real substrate of tyrosinase, with low catalytic efficiency and, therefore, delays the formation of d‐melanin

The research on tyrosinase inhibition and biological activity of cinnamaldehyde and its derivatives

酪氨酸酶（EC1.14.18.1）是一种含铜的氧化还原酶，广泛分布于微生物，动物，植物和人体中，是生物体生命活动的关键酶。若人体中酪氨酸酶异常表达能引起雀斑和黑色素沉积等皮肤病，且其在昆虫的蜕皮及果蔬褐变中起了关键的作用，因此酪氨酸酶抑制剂的研究引起了广泛的关注。肉桂醛主要存在于天然樟科植物桂皮中，在农业、医药上具有重要的作用。 本文首先以蘑菇酪氨酸酶为对象，研究了肉桂醛及其α位C上取代的三种衍生物：α-溴代肉桂醛、α-氯代肉桂醛和α-甲基肉桂醛对酶的抑制作用，结果表明四者抑制50%单酚酶活性的浓度（IC50）分别为1.210，0.075，0.140，0.440mmol/L；对二酚酶的IC5...Tyrosinase (EC 1.14.18.1), a copper containing enzyme, is widely distributed in microorganisms, animals, plants and human beings. It is the key enzyme in the vital function of biology. In human, tyrosinase is responsible for skin pigmentation abnormalities, such as flecks and defects. Furthermore, tyrosinase causes browning in vegetables, fruits and mushrooms. Therefore, The inhibitors of tyrosina...学位：理学硕士院系专业：生命科学学院_生物化学与分子生物学学号：2162013115257

Production of commercially suitable pectin methylesterase and polyphenol oxidase from agro-industrial wastes

Thesis(Master)--Izmir Institute of Technology, Food Engineering, Izmir, 2004Includes bibliographical references (leaves: 68)Text in English; Abstract: Turkish and Englishxiii, 80 leavesIn this study, some simple and effective extraction and/or partial purification procedures were developed to obtain pectin methylesterase (PME) and polyphenol oxidase (PPO) enzymes from orange peels and mushroom stems, respectively. Also, some characteristics of enzymes were investigated and their stable preparations were obtained in liquid or lyophilized forms. Valencia orange peels contain considerable PME activity (300-350 mL NaOH/min/100 g) that is quite stable during season for at least 5 months. The enzyme was ionically bound to cell walls and can not be extracted by homogenization with water. However, the addition of suitable amounts of NaCl (10 g /100 g extraction mixture) to pellet, obtained by homogenization of peels several times with water, and 30 min mixing (at 200 rpm) may be effectively used to extract the enzyme. The PME in orange peels contains almost the same amount of heat stable and heat labile fractions and the enzyme can not be activated by mild heating. A slight activation (almost 20 %) may be achieved by adding 1 mM CaCl2 to enzyme extracts. However, at higher concentrations the addition of CaCl2 was inhibitory. The PME activity in extracts, stabilized by use of 0.1 % Na-benzoate and 0.1 % K-sorbate, is stable almost 5 months at + 4 oC (maintains > 90 % of its activity). Thus, the commercial preparations of enzyme may be obtained in liquid form. The extracted PME was successfully used to prepare edible films from citrus pectin For the extraction of PPO, on the other hand, mushroom stems were first processed to acetone powder. The acetone powders were then extracted with Na-phosphate buffer and partially purified with ammonium sulfate (90 % saturation) or acetone precipitation (2-fold). Following dialysis, the recoveries and purification folds obtained from the partial purification of monophenolase activity of PPO from the same acetone powder were 74-86 % and 3.4-4.3 and 55-67 % and 5.4-6.2 for ammonium sulfate and acetone precipitations, respectively. Thus, it appears that the ammonium sulfate precipitation gives a higher yield but lower purity. The monophenolase activity of partially purified PPO is heat labile and showed inactivation above 45 oC. The enzyme exhibited a pH optimum between pH 6.0 and 8.0. The pH stability of enzyme was maximal at pH 7.0 and 8.0. However, at pH 4.0 the enzyme lost most of its activity after 24 h incubation. The optimum temperature of enzyme was found as 40 C. The monophenolase activity of PPO enzyme showed no stability in acetone powders at + 4 oC. However, it showed good stability at -18 oC for two months with retention of 60-70 % of its activity. The PPO partially purified with ammonium sulfate precipitation and dialysis, and lyophilized by using dextran or saccharose as supporting materials also retained its monophenolase and diphenolase activities for three months at -18 oC. The effect of lyophilization with dextran on temperature stability of enzyme was insignificant. However, lyophilization with dextran reduced the pH stability of monophenolase activity at 4.0 moderately. In addition to its monophenolase activity on tyrosine and diphenolase activity on L-DOPA, PPO lyophilized with dextran can also use phloridzin as substrate. Thus, it appears that the enzyme may be used in different food applications including the production of antioxidants and colorants, modification of proteins, fermentation of cocoa and black tea, etc

Tyrosinase inhibitory activity of native plants from central Argentina: Isolation of an active principle from Lithrea molleoides

Screening of 91 native plants from central Argentina was carried out with the aim of finding new sources of anti-tyrosinase compounds. Extracts obtained from Achyrocline satureioides, Artemisia verlotiorum, Cotoneaster glaucophylla, Dalea elegans, Flourensia campestris, Jodina rhombifolia, Kageneckia lanceolata, Lepechinia floribunda, Lepechinia meyenii, Lithrea molleoides, Porlieria microphylla, Pterocaulon alopecuroides, Ruprechtia apetala, Senna aphylla, Sida rhombifolia, Solanum argentinum, Tagetes minuta and Thalictrum decipiens exhibited more than 90% inhibition of tyrosinase monophenolase activity at 1000 μg ml-1. D. elegans, L. meyenii and L. molleoides were the most potent with IC50 values of 0.48, 10.43 and 3.77 μg ml-1, respectively. D. elegans, L. molleoides and T. decipiens also showed more than 90% inhibition of diphenolase activity at 1000 μg ml-1, with the first of these being the most effective (IC50 = 49.27 μg ml-1). (Z,Z)-5-(trideca-4,7-dienyl)-resorcinol (1) was isolated from L. molleoides as an effective tyrosinase inhibitor with l-tyrosine or l-DOPA as substrates (IC50 = 0.49 and 14.94 μg ml-1, respectively). Compound 1 was 37 times more active in monophenolase inhibitory activity than kojic acid used as a reference. Effective extracts as well as (Z,Z)-5-(trideca-4,7-dienyl)-resorcinol could prove to be promising preservative agents for use in the food industry.Fil: Chiari, Maria Eugenia. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Joray, Mariana Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentina. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; ArgentinaFil: Ruiz, Gustavo Miguel. Universidad Católica de Córdoba. Facultad de Ciencias Agropecuarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Palacios, Sara Maria. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Carpinella, Maria Cecilia. Universidad Católica de Córdoba. Facultad de Ciencias Químicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; Argentin