Quantitation of NAD+ biosynthesis from the salvage pathway in Saccharomyces cerevisiae

Nicotinamide adenine dinucleotide (NAD⁺) is synthesized via two major pathways in prokaryotic and eukaryotic systems: the de novo biosynthesis pathway from tryptophan precursors, or the salvage biosynthesis pathway from either extracellular nicotinic acid or various intracellular NAD⁺ decomposition products. NAD⁺ biosynthesis via the salvage pathway has been linked to an increase in yeast replicative lifespan under calorie restriction (CR). However, the relative contribution of each pathway to NAD⁺ biosynthesis under both normal and CR conditions is not known. Here, we have performed lifespan, NAD⁺ and NADH (the reduced form of NAD⁺) analyses on BY4742 wild‐type, NAD⁺ salvage pathway knockout (npt1Δ) and NAD⁺ de novo pathway knockout (qpt1Δ) yeast strains cultured in media containing either 2% glucose (normal growth) or 0.5% glucose (CR). We have utilized ¹⁴C labelled nicotinic acid in the culture media combined with HPLC speciation and both UV and ¹⁴C detection to quantitate the total amounts of NAD⁺ and NADH and the amounts derived from the salvage pathway. We observed that wild‐type and qpt1Δ yeast exclusively utilized extracellular nicotinic acid for NAD⁺ and NADH biosynthesis under both the 2% and 0.5% glucose growth conditions, suggesting that the de novo pathway plays little role if a functional salvage pathway is present. We also observed that NAD⁺ concentrations decreased in all three strains under CR. However, unlike the wild‐type strain, NADH concentrations did not decrease and NAD⁺: NADH ratios did not increase under CR for either knockout strain. Lifespan analyses revealed that CR resulted in a lifespan increase of approximately 25% for the wild‐type and qpt1Δ strains, while no increase in lifespan was observed for the npt1Δ strain. In combination, these data suggest that having a functional salvage pathway is required for lifespan extension under CR. Copyright © 2009 John Wiley & Sons, Ltd.     

A preliminary study of monosaccharide composition and α‐glucosidase inhibitory effect of polysaccharides from pumpkin (Cucurbita moschata) fruit

In this work, crude polysaccharide extracts were extracted from pumpkin (Cucurbita moschata) fruit by hot water. After removal of proteins and purification, polysaccharides of pumpkin fruit (PP1‐1) were subjected to structural identification. Gas chromatography analysis indicated that PP1‐1 comprised of galactose (86.4%), and glucose (13.6%). The molecular weight of PP1‐1 was measured to be 0.87 × 10⁴ Da by gel permeation chromatography. The inhibitory kinetic evaluation showed that it was non‐competitive inhibition of PP1‐1 on the α‐glucosidase‐catalysed hydrolysis of PNPG. The Michaelis–Menten constant (K_m) was 0.106 m , and the inhibitory constants (K_i), 0.435 mg.     

KINETIC STUDIES AND MECHANISM OF CATALYSIS OF MALIC DEHYDROGENASE FROM DIOSCOREA ROTUNDATA TUBER

ABSTRACT The kinetic mechanism of catalysis by malic dehydrogenase (EC 1.1.1.37) isolated from yam (Dioscorea rotundata) tuber has been delineated. Initial velocity studies with the enzyme in the presence and absence of products of the reaction revealed an ordered sequential mechanism. The K_m values obtained from secondary plots were 0.05, 0.08, 0.48 and 2.56 mM for NADH, OAA, and NAD⁺ and L-malic acid, respectively. Product inhibition studies in both the forward and backward reactions support an ordered Bi-Bi sequential mechanism. This begins with an obligatory binding of NAD⁺ to form the first binary complex and a final release of NADH.     

Enzymatic synthesis of ethyl hexanoate by transesterification

Summary Enzymatic synthesis of ethyl hexanoate by means of an acyl transfer reaction has been studied by using an immobilized Rhizomucor miehei lipase (RML). The effect of reaction parameters on ester synthesis has been investigated. Rhizomucor miehei lipase showed more specificity than other lipases when ethyl hexanoate was synthesized in n‐hexane. Maximum ester synthesis was obtained by using a 0.5 m substrate concentration (equimolar ratio). Temperatures in the range of 45–55 °C were found to be optimum and at higher temperatures (>60 °C) deactivation of enzyme was observed. Higher molar concentrations of hexanoic acid inhibited RML, but no inhibitory effect of ethyl caprate, even at higher molar concentrations, was observed. Apparent kinetic parameters have been determined. The values are as follows: K_M (ester), 0.0135 m ; K_M (acid), 0.08466; K_i (ester), 3.07 m ; K_i (acid), 0.550 m ; V_max, 1.861 µmol min^?1 mg^?1 enzyme.     

Inhibitory effects of selected pesticides on peroxidases purified by affinity chromatography

The objective of this study was to determine the in vitro inhibition effects of seven commonly used pesticides including 2,4-d-acid dimethylamine, fenoxaprop-p-ethyl, glyphosate isopropylamine, haloxyfop-p-methyl, cypermethrin, λ-cyhalothrin, and dichlorvos on the peroxidase purified from turnip (Brassica rapa L.) and black radish (Raphanus sativus L.) using 4-amino benzohydrazide affinity column chromatography. The purification factors for the turnip and black radish peroxidases were found to be 263.29-fold (with a yield of 12.89%) and 36.20-fold (with a yield of 6.90%), respectively. Among these compounds, λ-cyhalothrin showed the strongest inhibitory effect against turnip peroxidase (K_i: 1.23 × 10^?2 ± 0.21 × 10^?2 mM) as noncompetitive inhibition. On the other hand, cypermethrin demonstrated the highest inhibition effect against black radish peroxidase (K_i: 2.14 × 10^?2 ± 0.08 × 10^?2 mM) as competitive inhibition.     

Water‐forming NADH oxidase protects Torulopsis glabrata against hyperosmotic stress

A heterologous water‐forming NADH oxidase was introduced into Torulopsis glabrata and the effect on cell growth under hyperosmotic conditions was investigated. Expression of the noxE gene from Lactococcus lactis NZ9000 in T. glabrata resulted in a marked decrease in the NADH : NAD⁺ ratio and higher activities of key enzymes in water‐regenerating pathways, leading to an increase in intracellular water content. NaCl‐induced reactive oxygen species production was also decreased by the introduction of NADH oxidase, resulting in a significant increase in the growth of T. glabrata under hyperosmotic stress conditions (3824 mOsmol/kg). The results indicated that the osmotolerance of cells can be enhanced by manipulating water‐production pathways. Copyright © 2009 John Wiley & Sons, Ltd.     

Carotenoids in yellow‐ and red‐fleshed papaya (Carica papaya L)

Vitamin A deficiency is a disorder of public health importance in Sri Lanka. A recent national survey revealed that 36% of preschool children in Sri Lanka have vitamin A deficiency (serum retinol <0.2 µg ml^?1). In view of its well‐established association with child morbidity and mortality, this is a reason for concern. One of the main fruits which has been recommended for prevention of vitamin A deficiency in Sri Lanka is papaya (Carica papaya L). In this study the carotenoid profiles of yellow‐ and red‐fleshed papaya were analysed by medium‐pressure liquid chromatography (MPLC) and UV‐vis spectrophotometry. A section of yellow‐fleshed papaya showed small carotenoid globules dispersed all over the cell, whereas in red‐fleshed papaya the carotenoids were accumulated in one large globule. The major carotenoids of yellow‐fleshed papaya were the provitamin A carotenoids β‐carotene (1.4 ± 0.4 µg g^?1 dry weight (DW)) and β‐cryptoxanthin (15.4 ± 3.3 µg g^?1 DW) and the non‐provitamin A carotenoid ζ‐carotene (15.1 ± 3.4 µg g^?1 DW), corresponding theoretically to 1516 ± 342 µg kg^?1 DW mean retinol equivalent (RE). Red‐fleshed papaya contained the provitamin A carotenoids β‐carotene (7.0 ± 0.7 µg g^?1 DW), β‐cryptoxanthin (16.9 ± 2.9 µg g^?1 DW) and β‐carotene‐5,6‐epoxide (2.9 ± 0.6 µg g^?1 DW), and the non‐provitamin A carotenoids lycopene (11.5 ± 1.8 µg g^?1 DW) and ζ‐carotene (9.9 ± 1.1 µg g^?1 DW), corresponding theoretically to 2815 ± 305 µg kg^?1 DW mean RE. Thus the carotenoid profile and organisation of carotenoids in the cell differ in the two varieties of papaya. This study demonstrates that carotenoids can be successfully separated, identified and quantified using the novel technique of MPLC. Copyright © 2003 Society of Chemical Industry     

Improving protein secretion of a transglutaminase-secreting Corynebacterium glutamicum recombinant strain on the basis of 13C metabolic flux analysis

Corynebacterium glutamicum is known as a host species for amino acid production. This microorganism was recently noticed as a host that produces secreted proteins. In this study, we performed ¹³C metabolic flux analysis (¹³C-MFA) on a recombinant C. glutamicum strain that secretes a heterologous transglutaminase (TGase) to improve TGase secretion. For the ¹³C-MFA of a TGase-secreting C. glutamicum strain in batch cultivation, a ¹³C-labeling experiment and measurement of mass isotopomer distributions of proteinogenic amino acids were performed, and metabolic fluxes were determined considering the changes in fractional ¹³C-labeling of proteinogenic amino acids with respect to culture time. The TGase yield increased at the stationary phase but decreased toward its end. The results of ¹³C-MFA revealed that the flux from glycolysis to the TCA cycle gradually increased during TGase secretion. We speculate that the NADH/NAD⁺ ratio in the cells increases and that as a result, the specific glucose uptake rate decreases in the stationary phase because of the increased flux of the TCA cycle. Since it is expected that a decrease in the NADH/NAD⁺ ratio would improve the TGase yield, we tried to enhance lactate production in a TGase-secreting C. glutamicum strain to decrease cellular NADH levels by increasing the pH level in the culture. The TGase yield increased in 1.4-fold by increasing the pH from 6.7 to 7.2, indicating that the TGase yield was successfully improved on the basis of the ¹³C-MFA.     

Antioxidant and semicarbazide‐sensitive amine oxidase inhibitory activities of alginic acid hydroxamates

The commercial polysaccharides of alginic acid (medium (3500 cps, 2% solution) and low (250 cps, 2% solution) viscosities) were esterified with acidic methanol (1 mmol L^?1 HCl) at 4 °C with gentle stirring for 5 days to obtain methyl esters of medium‐viscosity alginic acid (ME‐MVA) and low‐viscosity alginic acid (ME‐LVA). These ME‐MVA and ME‐LVA were reacted with alkaline hydroxylamine to obtain medium‐viscosity alginic acid hydroxamates (MVA‐NHOH) and LVA‐NHOH. The percentages of hydroxamic acid content in MVA‐NHOH and LVA‐NHOH were calculated as 25% and 20%, respectively. The hydroxamate derivatives of alginic acid were used to test the antioxidant and semicarbazide‐sensitive amine oxidase (SSAO) inhibitory activities in comparison with original materials (MVA and LVA). The half‐inhibition concentrations, IC₅₀, of scavenging activity against 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) were 24.5 and 29.8 µg mL^?1 for MVA‐NHOH and LVA‐NHOH, respectively. However, few scavenging activities of the MVA and LVA were found at the same concentrations. The IC₅₀ of the positive control of butylated hydroxytoluene was 5 µg mL^?1. The scavenging activity of DPPH radical was pH‐dependent, and the optimal pH for both of MVA‐NHOH and LVA‐NHOH was the Tris‐HCl buffer (pH 7.9). Using electron spin resonance (ESR) to detect the activity of scavenging hydroxyl radicals, both alginic acid hydroxamates showed dose‐dependent scavenging activities, and the IC₅₀ was 90 and 92 µg mL^?1, respectively, for MVA‐NHOH and LVA‐NHOH. Both alginic acid hydroxamates also exhibited protection against hydroxyl radical‐mediated DNA damage. Both MVA‐NHOH and LVA‐NHOH showed dose‐dependent inhibitory activities against bovine SSAO (2.53 units); the IC₅₀ was 0.16 and 0.09 µg mL^?1, respectively, for MVA‐NHOH and LVA‐NHOH, compared with 3.81 µg mL^?1 of semicarbazide (positive controls). Amine oxidase activity staining also revealed that both MVA‐NHOH and LVA‐NHOH exhibited SSAO inhibitory activities. Both MVA‐NHOH and LVA‐NHOH showed mixed non‐competitive inhibition against bovine SSAO. It was found that the V′_max value was reduced and the K′_m value was either increased (added MVA‐NHOH, 0.05 µg mL^?1) or reduced (added LVA‐NHOH, 0.11 µg mL^?1) in the presence of alginic acid hydroxamate. Copyright © 2006 Society of Chemical Industry     

Polyphenol oxidase properties,anti-urease,and anti-acetylcholinesterase activity of Diospyros lotus L. (Plum Persimmon)

Diospyros lotus fruit polyphenol oxidase was purified using affinity chromatography, resulting in a 15-fold enrichment in specific activity. The purified enzyme, having 16.5 kDa molecular weight on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, exhibited the highest activity toward 4-methylcatechol. Maximum diphenolase activity was reached at pH 7.0 and 60°C in the presence of 4-methylcatechol. K_m and V_max values were calculated as 3.8 mM and 1250 U/mg protein, respectively. Ascorbic acid was a promising inhibitor with an IC₅₀ value of 0.121 µM. The activity of the purified enzyme was stimulated by Fe²⁺, Sr²⁺, Zn²⁺, and K⁺ and deeply inhibited by Hg²⁺, at 1 mM final concentration. Aqueous extract of Diospyros lotus L. fruit showed strong substantial urease and acetylcholinesterase inhibition, with IC₅₀ values of 1.55 ± 0.05 and 16.75 ± 0.11 mg/mL, respectively.     

Growth of a tropical marine yeast Yarrowia lipolytica NCIM 3589 on bromoalkanes: relevance of cell size and cell surface properties

Yarrowia lipolytica 3589, a tropical marine yeast, grew aerobically on a broad range of bromoalkanes varying in carbon chain length and differing in degree and position of bromide group. Amongst the bromoalkanes studied, viz. 2‐bromopropane (2‐BP), 1‐bromobutane (1‐BB), 1,5‐dibromopentane (1,5‐DBP) and 1‐bromodecane (1‐BD), the best utilized was 1‐BD, with a maximal growth rate (μ_max) of 0.055 h^?1 and an affinity ratio (μ_max/K_s) of 0.022. Utilization of these bromoalkanes as growth substrates was associated with a concomitant release of bromide (8202.9 µm ) and cell mass (36 × 10⁹ cells/ml), occurring maximally on 1‐BD. Adherence of yeast cells to these hydrophobic bromoalkanes was observed microscopically, with an increase in cell size and surface hydrophobicity. The maximal cell diameter was for 1‐BD (4.66 µm), resulting in an increase in the calculated cell surface area (68.19 µm²) and sedimentation velocity (1.31 µm/s). Cell surface hydrophobicity values by microbial adhesion to solvents (MATS) analysis for yeasts grown on bromoalkanes and glucose were significantly high, i.e. >80%. Similarly, water contact angles also indicate that the cell surface of yeast cells grown in glucose possess a relatively more hydrophilic cell surface (θ = 49.1°), whereas cells grown in 1‐BD possess a more hydrophobic cell surface (θ = 90.7°). No significant change in emulsification activity or surface tension was detected in the cell‐free supernatant. Thus adherence to the bromoalkane droplets by an increase in cell size and surface hydrophobicity leading to debromination of the substrate might be the strategy employed in bromoalkane utilization and growth by Y. lipolytica 3589. Copyright © 2011 John Wiley & Sons, Ltd.     

The role of K+ and H+ transport systems during glucose‐ and H2O2‐induced cell death in Saccharomyces cerevisiae

Glucose, in the absence of additional nutrients, induces programmed cell death in yeast. This phenomenon is independent of yeast metacaspase (Mca1/Yca1) and of calcineurin, requires ROS production and it is concomitant with loss of cellular K⁺ and vacuolar collapse. K⁺ is a key nutrient protecting the cells and this effect depends on the Trk1 uptake system and is associated with reduced ROS production. Mutants with decreased activity of plasma membrane H⁺‐ATPase are more tolerant to glucose‐induced cell death and exhibit less ROS production. A triple mutant ena1‐4 tok1 nha1, devoid of K⁺ efflux systems, is more tolerant to both glucose‐ and H₂O₂‐induced cell death. We hypothesize that ROS production, activated by glucose and H⁺‐ATPase and inhibited by K⁺ uptake, triggers leakage of K⁺, a process favoured by K⁺ efflux systems. Loss of cytosolic K⁺ probably causes osmotic lysis of vacuoles. The nature of the ROS‐producing system sensitive to K⁺ and H⁺ transport is unknown. Copyright © 2010 John Wiley & Sons, Ltd.     

Gastric H+, K+-ATPase Inhibition,and Antioxidant Properties of Selected Commonly Consumed Vegetable Sources

Oxidative stress and upregulation of gastric H⁺, K⁺-ATPase enzyme activity have been known to cause ulcer pathogenicity for which safer drugs are yet to be identified. Aqueous extracts of seven commonly consumed vegetable sources were screened for inhibition of H⁺, K⁺-ATPase and antioxidant activities. Results indicated that Z. officinale (Ginger) followed by M. arvensis (Pudina) are potent gastroprotective sources with inhibition of H⁺, K⁺-ATPase of IC₅₀ of 18.3 ± 0.7 and 25.2 ± 0.9 μg gallic acid equivalents/ml respectively, which is almost equivalent or better than the known inhibition of H⁺, K⁺-ATPase—Omeprazole (IC₅₀ ?27 μg/ml). Further, all these vegetable extracts showed multi-potent antioxidant activity, such as free radical scavenging, reducing power ability, and inhibition of lipid peroxidation, which are required to inhibit complex steps of ulcerations. On the basis of the absolute amounts and potency of inhibition of H⁺, K⁺-ATPase as well as antioxidant activity of individual phenolic acids, the relative percentage contribution of phenolic acids from different vegetable extracts to both inhibition of H⁺, K⁺-ATPase and antioxidant activity was calculated and data revealed that gentisic and protocatechuic acid contributes significantly to both inhibition of H⁺, K⁺-ATPase and antioxidant activity.     

Higher bioavailability of isoflavones after a single ingestion of aglycone-rich fermented soybeans compared with glucoside-rich non-fermented soybeans in Japanese postmenopausal women

BACKGROUND: There have been conflicting study results concerning how the food matrix affects the bioavailability of isoflavone aglycone and glucoside. In this study the bioavailability of isoflavones after a single ingestion of aglycone‐rich fermented soybeans (Fsoy) and glucoside‐rich non‐fermented soybeans (Soy) was compared. Eleven healthy postmenopausal Japanese women were recruited for a randomised, double‐blind, crossover trial and consumed Fsoy or Soy powder dissolved in hot water. Blood samples were collected 0, 1, 2, 3, 4, 6, 8, 12 and 24 h and urine samples from 0 to 48 h after ingestion of the powders. The Fsoy and Soy powders ingested had the same total isoflavone content (95 µmol), but the former was rich in aglycone (90.6 µmol) while the latter was rich in glucoside (81.9 µmol). RESULTS: Serum concentrations of total isoflavones after 1–4 h were significantly higher in the Fsoy group than in the Soy group. The Fsoy group showed significantly higher maximum concentration (Cmax: 2.79 ± 0.13 vs 1.74 ± 0.13 µmol L^?1) and area under the curve (AUC_{0–24 h}: 23.78 ± 2.41 vs 19.95 ± 2.03 µmol day L^?1) and lower maximum concentration time (Tmax: 1.00 ± 0.00 vs 5.00 ± 0.67 h) compared with the Soy group. The cumulative urinary excretion of total isoflavones after 2 h was significantly higher in the Fsoy group than in the Soy group. Individual isoflavones (daidzein, genistein and glycitein) showed similar trends to total isoflavones. Equol (a metabolite from daidzein) did not differ between the two groups. CONCLUSION: The results of this study demonstrated that the isoflavones of aglycone‐rich Fsoy were absorbed faster and in greater amounts than those of glucoside‐rich Soy in postmenopausal Japanese women. Copyright © 2010 Society of Chemical Industry     

REGULATION OF PHENYLALANINE AMMONIA-LYASE ENZYME IN ANNONA FRUIT: KINETIC CHARACTERISTICS AND INHIBITORY EFFECT OF AMMONIA

In this work, we analyzed the kinetic properties of phenylalanine ammonia‐lyase (PAL) extracted from “cherimoya” (Annona cherimola Mill.) fruits ripened at ambient temperature (20C) and stored under several environmental conditions, including high CO₂ levels (20%) and low temperature (6C). The effect of different ammonia‐related compounds on cherimoya PAL activity was also evaluated. PAL exhibited two different K_mvalues for L‐phenylalanine (L‐Phe ) and negative substrate cooperativity, with Hill coefficient (n_app) values reaching 0.64 and 0.71 for low temperature and high CO₂ levels, respectively. The kinetic analysis revealed that ammonia produced mixed inhibition of PAL enzyme, with inhibition constants (K_i and K_i′) values of 0.57 ± 0.2 mM and 2.54 ± 0.2 mM. We propose that the regulation of PAL by ammonia inhibition and the negative cooperativity may be essential in adjusting the active phenylpropanoid metabolism in Annonas to the requirement of L‐Phe and in consequence, to the carbon skeleton demand for other anabolic pathways.     

Latent polyphenol oxidase from quince fruit pulp (Cydonia oblonga): purification,activation and some properties

Quince fruit polyphenol oxidase (PPO) was partially purified using a combination of phase partitioning in Triton X‐114 and PEG 8000/phosphate with a final ammonium sulfate fractionation between 30% and 75%, to avoid the deep browning of the enzyme due to the high amount of oxidizing substances present in the quince pulp. The clean and stable enzyme was partially purified in a latent form and could be optimally activated by the presence of 0.5 g dm^?3 sodium dodecyl sulfate (SDS) with an optimum pH of 5.0. In the absence of SDS, the enzyme showed maximum activity at acid pH. The apparent kinetic parameters of the latent enzyme were determined at pH 5.0, the V_m value being 15 times higher in the presence of SDS than in its absence, whereas the K_M was the same in both cases, with a value of 1.2 mmol L^?1. The effect of several inhibitors was studied, tropolone being the most active with a K_i value of 4.7 µmol L^?1. In addition, the effect of cyclodextrins was studied, and the complexation constant (K_c) between 4‐tert‐butylcatechol and cyclodextrins was calculated using an enzymatic method. The value obtained for K_c was 15 310 mol L^?1. Copyright © 2006 Society of Chemical Industry     

Effects of tea polyphenols on the activities of soybean trypsin inhibitors and trypsin

Tea polyphenols (TPs) and other materials were extracted from Chinese green tea, and their effects on trypsin inhibitors and trypsin were analysed. TPs were found to have a deactivation effect on both Kunitz trypsin inhibitor (KTI) and Bowman–Birk trypsin inhibitor (BBTI). KTI was more easily deactivated than BBTI by complexing with TPs. The deactivation effect of TPs on KTI and BBTI reached a maximum at a TP/KTI ratio of 25 and a TP/BBTI ratio of 16. However, the deactivation effect of TPs on KTI and BBTI was reduced dramatically when KTI and BBTI were already complexed with trypsin. TPs were also found to inhibit trypsin. The inhibitory activity of TPs, KTI and BBTI on trypsin was found to decrease in the order BBTI > gtTI > gtPs. Complete inhibition of trypsin by TPs could not be achieved. When the TP concentration was increased to about 17 µg ml^?1, the residual activity of trypsin was maintained at 400 TU mg^?1, equivalent to 32% of the initial trypsin activity. In TP inhibition the K_M value for trypsin remained unchanged at 5.88 × 10^?4 mol l^?1 and V_max decreased when benzoyl‐DL ‐arginine‐p‐nitroanilide (BAPNA) was used as substrate. The pattern of trypsin inhibition by TPs is non‐competitive. Copyright © 2004 Society of Chemical Industry     

Inhibition of polyphenol oxidase obtained from various sources by 2,3‐diaminopropionic acid

This paper reports for the first time the inhibition of the catecholase activities of mushroom, artichoke (Cynara scolymus L) and Ocimum basilicum L polyphenol oxidase by 2,3‐diaminopropionic acid. Polyphenol oxidases from artichoke and O basilicum L were purified by ammonium sulfate precipitation, dialysis and a Sepharose 4B‐L ‐tyrosine‐p‐aminobenzoic acid‐affinity column. In inhibition studies, 2,3‐diaminopropionic acid showed uncompetitive inhibition for mushroom PPO using catechol and pyrogallol as substrates, competitive inhibition for O basilicum L PPO using catechol as a substrate, and uncompetitive inhibition for artichoke PPO using catechol as a substrate. Furthermore, sodium azide, which is an inhibitor of PPO, was used as an inhibitor for comparison with the inhibition potency of 2,3‐diaminopropionic acid. The highest 2,3‐diaminopropionic acid inhibition observed with O basilicum L (K_i = 0.89 mM ), followed by artichoke (K_i = 1.42 mM ) and mushroom (K_i = 2.47 mM ), respectively. Copyright © 2005 Society of Chemical Industry     

Aflatoxins in composite spices collected from local markets of Karachi,Pakistan

This survey was carried out to evaluate the occurrence of total aflatoxins (AFs; B₁+B₂+G₁+G₂) in unpacked composite spices. A total of 75 samples of composite spices such as biryani, karhai, tikka, nihari and korma masalas were collected from local markets of Karachi, Pakistan, and analysed using HPLC technique. The results indicated that AFs were detected in 77% (n = 58) samples ranging from 0.68 to 25.74 µg kg^?1 with a mean of 4.63 ± 0.95 µg kg^?1. In 88% (n = 66) samples, AFs level was below the maximum limits (ML = 10 µg kg^?1) as imposed by EU. Furthermore, 61% (n = 46) tested samples contained AFs level between 1 and 10 µg kg^?1, 9% (n = 7) exhibited AFs contamination ranged 10?20 µg kg^?1 and only 3% (n = 2) of the investigated samples contained AFs levels higher than the ML of 20 µg kg^?1 for total aflatoxins as set by the USA. It was concluded that there is need to establish a strict and continuous national monitoring plan to improve safety and quality of spices in Pakistan.     

Characteristics of the leaf parts of some traditional Korean salad plants used for food

BACKGROUND: Total phenolics content, antioxidant activity and cytotoxicity of the methanol extracts from leaf parts of 13 Korean traditional salad plants were investigated in order to determine their properties. RESULTS: The highest phenolics content (mg ferulic acid equivalents kg^?1 dry weight (d.w.), omit one) was found in methanol extracts from Polygonum aviculare, at 293.7 ± 6.0, followed by Euonymus alatus, at 250.7 ± 3.3, Saxifraga stolonifera, at 125.0 ± 8.1 and Ligularia fischeri, at 122.5 ± 5.9. The methanol plant extracts dose‐dependently increased free radical scavenging activity. Methanol extracts of Polygonum aviculare, Euonymus alatus and Saxifraga stolonifera, at 31 mg kg^?1, exhibited the highest 1,1‐diphenyl‐2‐picrylhydrazyl (DPPH) radical scavenging activity (%) by 90.8 ± 4.2, 85.7 ± 3.9 and 64.1 ± 3.2, respectively. According to 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay, the methanol extracts from Portulaca oleracea (IC₅₀ < 25.0 µg mL^?1) showed the highest cytotoxicity against Calu‐6, followed by Plantago asiatica (49.2 µg mL^?1) and Osmunda japonica (89.6 µg mL^?1). CONCLUSION: Total phenolics content of the tested plant extracts was correlated with the DPPH radical scavenging activity, suggesting the phenolics compounds are contributing to the antioxidant properties of Korean salad plants. The leaf parts of the 13 Korean traditional salad plants described here that are currently used as foods may also provide some benefit to human health, and research into their potential benefits as preventative and/or therapeutic agents is warranted. Copyright © 2008 Society of Chemical Industry