p-HYDROXYPHENYLPROPIONIC ACID (PHPPA) AND 3,4-DIHYDROXYPHENYLPROPIONIC ACID (3,4-DPPA) AS SUBSTRATES FOR MUSHROOM TYROSINASE

p-Hydroxyphenylpropionic acid (PHPPA) and 3,4- dihydroxyphenylpropionic acid (3,4-DPPA) serve as substrates for tyrosinase. The Km value of 3,4-DPPA for tyrosinase is 1.3 mM. The yellow o-quinone of 3,4-DPPA (4-carboxyethyl-o-benzoquinone) (λ_max= 400nm), is detected initially and it is then converted to a red product(s) (λ_max= 480±10 nm), the o-quinone of 6,7-dihydroxy 3-dihydrocumarin (dihydroesculetin). When the concentration of the latter is relatively high, it polymerizes to a final brown product(s), characterized by an ill-defined spectrum.
H₂O₂ shortens the lag period of PHPPA hydroxylation, hastens the conversion of the yellow o-quinone of 3,4-DPPA to the red o-quinone of dihydroesculetin, and prevents the polymerization of the latter to the final brown product(s).
The relatively unstable o-quinone of 3,4-DPPA interacts with amines such as hydroxylamine (NH₂OH), p-aminosalicylic acid (PASA) and p-aminobenzoic acid (PABA), forming relatively stable final product(s) characterized by different spectra from those formed in their absence.
Acetohydroxamic acid (AHA) and salicylhydroxamic acid (SHAM) each has an effect on the spectrum of product(s) obtained when 3,4-DPPA is oxidized by tyrosinase, indicating that these hydroxamic acids derivatives interact with the o-quinone of 3,4-DPPA. The spectrum of the final product(s) was also different when 3,4-DPPA was oxidized by tyrosinase in the presence of benzenesulfinic acid than in its absence, suggesting the formation of a stable phenylsulfonyl derivative.     

p-HYDROXYPHENYLACETIC ACID AND-3,4-DIHYDROXYPHENYLACETIC ACID AS SUBSTRATES FOR MUSHROOM TYROSINASE

The hydroxylation of p-hydroxyphenylacetic acid (PHPAC) and the oxidation of 3,4-dihydroxyphenylacetic acid (DOPAC) by mushroom tyrosinase are illustrated. DOPAC quinone (4-carboxy-methyl-o-benzoquinone (λ_max= 400±10 nm) is the initial pigmented product formed when DOPAC is oxidized by the enzyme. DOPAC quinone is very unstable, and, once formed, is converted rapidly to further oxidation product(s).
The relationships between the rate of p-dihydroxyphenylacetic acid hydroxylation and 3,4-dihydroxyphenylacetic acid oxidation as a function of various concentrations of each substrate and of mushroom tyrosinase, are described.
The effect of the addition of various chemicals that can potentially conjugate otherwise affect DOPAC quinone was studied The Km value of 3,4-dihydroxyphenylacetic acid for mushroom tyrosinase was estimated to be 4.0 mM.     

Browning prevention by ascorbic acid and 4-hexylresorcinol: different mechanisms of action on polyphenol oxidase in the presence and in the absence of substrates

ABSTRACT:  We have investigated the mechanism of action of 4-hexylresorcinol (4-HR) and ascorbic acid (AA) on the polyphenol oxidase (PPO) catalyzed oxidation of phenolic substrates. Incubation of PPO with 4-HR diminishes strongly PPO activity. This effect can be erroneously interpreted, due to the high affinity of 4-HR for PPO, as irreversible inactivation of PPO. However, PPO activity can be recovered by dialysis after incubation with 4-HR. 4-hexylresorcinol is a canonical enzyme inhibitor that binds preferentially to the oxy form of PPO. It is a mixed-type inhibitor, because it influences both apparent V _max (1.26 compared with 0.4 units in the absence and presence of 4-HR, respectively) and K _m values (0.28 mM compared with 0.97 mM in the absence and in the presence of 4-HR, respectively) of PPO. AA can prevent browning by 2 different mechanisms: In the absence of PPO substrates it inactivates PPO irreversibly, probably through binding to its active site, preferentially in its oxy form. In the presence of PPO substrates, AA reduces PPO oxidized reaction products, which results in a lag phase when measuring PPO activity by monitoring dark product formation but not when monitoring O₂ consumption. The simultaneous use of both 4-HR and AA on PPO results in additive prevention of browning.     

Reaction Between Malvidin 3-Glucoside and (+)-Catechin in Model Solutions Containing Different Aldehydes

ABSTRACT: The contribution of the aldehyde composition of wine spirit to the color changes in Port red wine was studied in model solutions. Malvidin 3-glucoside was shown to be very reactive towards catechin in the presence of different aldehydes: acetaldehyde, isovaleraldehyde, benzaldehyde, propionaldehyde, isobutyraldehyde, formaldehyde, and 2-methylbutyraldehyde. LC/MS data confirmed the formation of oligomeric pigments resulting from the reaction between the anthocyanin and the flavanol (colored products) and between 2 flavanol units (colorless products) mediated by each aldehyde assayed. The UV-visible spectra of the colored pigments showed a λ_max bathochromically shifted relatively to the λ_max of original anthocyanins. All samples revealed a "blueing" and "darkening" color effects using the CIELAB system.     

Myricetin,an antioxidant flavonol,is a substrate of polyphenol oxidase

The present study demonstrates the antiradical efficiency of myricetin, a flavonol widely distributed in fruits and vegetables, by testing its ability to react with two different free radicals, ABTS^+· and DPPH^·. The polyphenolic nature of myricetin led us to consider the possibility of its oxidation by polyphenol oxidase (PPO). The results reported show that myricetin can be oxidised by PPO extracted and partially purified from broad bean seeds. The reaction was followed by recording spectral changes with time, maximal spectral changes being observed at 372 nm. The presence of two isosbestic points (at 274 and 314 nm) suggested that only one absorbing product was formed. The spectral changes were not observed in the absence of PPO. The oxidation rate varied with the pH, reaching its highest value at pH 5.5. The myricetin oxidation rate increased in the presence of SDS, an activing agent of polyphenol oxidase. Maximal activity was obtained at 1.3 mM SDS. The kinetic parameters were also determined: V _m = 1.35 µM min⁻¹, K _m = 0.3,mM , V _m/ K _m = 4.5 × 10⁻³ min⁻¹. Flavonol oxidation was inhibited by a selective PPO inhibitor such as cinnamic acid (K_I = 1 mM ). The results reported show that myricetin oxidation was strictly dependent on the presence of polyphenol oxidase. © 1999 Society of Chemical Industry     

Continuous Spectrophotometric Method for Determining Monophenolase and Diphenolase Activities of Pear Polyphenoloxidase

A continuous spectrophotometric method was based on the coupling reaction between 3-methyl-2-benzothiazolinone hydrazone (MBTH) and the quinone product of the oxidation of p-hydroxyphenyl propionic acid (PHPPA) or 3,4-dihydroxyphenyl propionic acid (DHPPA) in the presence of polyphenol oxidase. The monophenolase activity of pear PPO was characterized for the first time. Solubility and stability of the adduct formed at the optimum pH (4.3) enabled the system to reach steady-state, making it possible to determine monophenolase activity with short lag periods. This, together with the value of ε for the MBTH-quinone adduct, makes this method more sensitive than other continuous methods for assaying monophenolase and diphenolase activities.     

EFFECT OF SODIUM LAURYL SULFATE IN SOLUBILIZING PROTEIN FOR TRYPTOPHAN DETERMINATION

The solubilization of proteins in corn and sorghum, using sodium lauryl sulfate (NaLS) in a basic medium, was used for tryptophan determination. The experimental design evaluated different levels of sodium lauryl sulfate and sodium hydroxide, at various temperatures and times. Prior to treatment, the samples were dried, ground and defatted. These were hydrolyzed and tryptophan was quantified spectrophotometrically (λ_max= 565 nm). Casein was used as control and DL-tryptophan was used for generating the standard curve according to Beer's Law. From the results obtained, it was possible to conclude that sodium lauryl sulfate was very effective in solubilizing corn and sorghum protein for subsequent tryptophan determination.     

Thermal Conductivity of Pear, Sweet-cherry, Apricot, and Cherry-plum Juices as a Function of Temperature and Concentration

ABSTRACT:  The thermal conductivity of 4 fruit (pear, sweet-cherry, apricot, and cherry-plum) juices was measured with a coaxial-cylinder (steady-state) technique. Measurements were made, temperature range 20 to 120 °C, at concentrations between 12.2 and 50 °Brix. The uncertainty of the thermal conductivity measurements was estimated to be less than 2%. A semitheoretical method for the prediction of thermal conductivity of juices was proposed. It was found that the prediction Model IV,  λ ( T , x )/λ ( T ₀, x ) = ( T / T ₀) ⁿ   , where  λ  is the thermal conductivity, x is the concentration, and T is the temperature, developed in this work can be adopted with satisfaction. The thermal conductivity of juices can be predicted just by knowing the thermal conductivity  λ₀  at a reference temperature   T ₀  .     

PROPERTIES OF IAA OXIDASE FROM RIPENING TOMATO FRUIT

An indoleacetic acid (IAA) oxidizing enzyme was isolated from the pericarp of ripening tomato fruit by column chromatography fractionation. IAA oxidase from extracts of tomato pericarp exists in a high molecular weight form (>200,000) or in a low molecular weight form (∼40,000) or both depending upon extraction procedures. When IAA was oxidized by the high molecular weight form of IAA oxidase, a relatively long lag time was observed. The low molecular weight form of IAA oxidase activity showed no lag or a very short, 1- to 2-minute lag period. Enzyme activity was optimal at pH 6.1 and was stimulated by 2,4-dichlorophenol (DCP) and MnCl_2. Neither 3-methyleneoxindole nor indolealdehyde were found as a major IAA oxidation products. Moderate increase in activity of IAA oxidase activity was observed during ripening of tomato fruit.     

Development of a Predictive Model for Spoilage of Cooked Cured Meat Products and Its Validation Under Constant and Dynamic Temperature Storage Conditions

ABSTRACT:  In the present study, the spoilage flora of a sliced cooked cured meat product was studied to determine the specific spoilage organism (SSO). The physicochemical changes of the product during its storage in a temperature range of 0 to 12 °C were also studied. Among the primary models used to model the temperature effect on SSO growth, the modified Gompertz described better the experimental data than modified logistic and Baranyi. The derived growth kinetic parameters, such as maximum specific growth rate (μ_max) and lag phase duration (LPD), were modeled by using the square root and Arrhenius equation (secondary models). The latter described better the data of μ_max and LPD; therefore, this model was chosen for correlating temperature with kinetic parameters. The selection of the best model (primary or secondary) was based on some statistical indices (the root mean square error of residuals of the model, the coefficient of multiple determination, the F -test, the goodness of fit, the bias, and accuracy factor). The validation of the developed model was carried out under constant and dynamic temperature storage conditions. To validate its usefulness to similar products, another sliced cooked cured meat product stored under constant temperature conditions was also used. The log shelf life model was used for shelf life predictions based on the evident (visual defects) or the incipient spoilage (attainment of a certain spoilage level by SSO and/or chemical spoilage index). The possibility for shelf life predictions constitutes a valuable information source for the quality assurance systems of meat industries.     

Combined Carbon Dioxide and High Pressure Inactivation of Pectin Methylesterase, Polyphenol Oxidase, Lactobacillus plantarum and Escherichia coli

ABSTRACT: High pressure processing (HPP) and CO₂have both been shown to increase food product shelf-life. CO₂ was added at approximately 0.2 molar % to solutions processed at 500 to 800 MPa in order to further inactivate pectin methylesterase (PME), polyphenol oxidase (PPO), L. plantarum ATCC 8014, and E. coli K12. An interaction was found between CO₂ and pressure at 25 °C and 50 °C for PME and PPO, respectively. Activity of PPO was decreased by CO₂ at all pressure treatments. The interaction between CO₂ and pressure was significant for L. plantarum with a significant decrease in survivors due to the addition of CO₂ at all pressures studied. No significant effect on E. coli survivors was seen with CO₂ addition.     

Formation of biopolymer-coated liposomes by electrostatic deposition of chitosan

ABSTRACT:  The purpose of this study was to prepare stable biopolymer-coated liposome suspensions using an electrostatic deposition method. Liposome suspensions were produced by homogenizing 1% soy lecithin in acetate buffer (0.1 M, pH 3). Cationic chitosan (Mw approximately 200 kDa) solutions were mixed with anionic liposome suspensions ( d approximately 100 and 200 nm), and the effect of phospholipid concentration, chitosan concentration, and liposome size on the properties of the particles formed was determined. The particle size and charge (ζ-potential) were measured using dynamic light scattering and particle electrophoresis. The particle charge changed from –38 mV in the absence of chitosan to +60 mV in the presence of chitosan, indicating complex formation between the anionic liposomes and cationic chitosan molecules. Below a minimum critical chitosan concentration ( c _min), large aggregates were formed that phase separated within minutes, whose origin was attributed to formation of coacervates. On the other hand, above a maximum critical chitosan concentration ( c _max), large flocs were formed that sedimented within hours, whose formation was attributed to depletion flocculation. Minimum and maximum critical chitosan concentrations depended on liposomal concentration and size. At c _min < c < c _max, chitosan-coated liposomes were formed that did not aggregate and were stable to sedimentation. Coated liposomes had better stability to aggregation than uncoated liposomes when stored at ambient temperatures for 45 d. This study indicates that chitosan can be used to form biopolymer-coated liposomes with enhanced stability over uncoated liposomes.     

Viscous Food Matrix Influences Absorption and Excretion but Not Metabolism of Blackcurrant Anthocyanins in Rats

ABSTRACT:  The aim of the present study was to investigate the effect of a simultaneous intake of food and anthocyanins (ACNs) on ACN absorption, metabolism, and excretion. Blackcurrant ACNs (BcACNs) were dissolved in water with or without the addition of oatmeal and orally administered to rats, providing approximately 250 mg total ACNs per kilogram BW. Blood, urine, digesta, and tissue samples of the stomach, jejunum, and colon were subsequently collected at 0.25, 0.5, 1, 2, 3, 7, and 24 h. Identification and quantification of ACNs were carried out by Reversed phase-high-performance liquid chromatography (RP-HPLC) and liquid chromatography-mass spectrometry (LC-MS). Four major ACNs were present in the blackcurrant extract: delphinidin 3- O -glucoside, delphinidin 3- O -rutinoside, cyanidin 3- O -glucoside, and cyanidin 3- O -rutinoside. In plasma, the 4 ACNs of blackcurrant were identified and quantified. The time to reach maximal total ACN plasma concentration ( C _max BcACN/water = 0.37 ± 0.07 μmol/L; C _max BcACN/oatmeal = 0.20 ± 0.05 μmol/L) occurred faster after BcACN/water ( t _max= 0.25 h), than after BcACN/oatmeal administration ( t _max= 1.0 h). In digesta and tissue samples, the 4 original blackcurrant ACNs were detected. The relative concentration of rutinosides in the digesta increased during their passage through the gastrointestinal tract, while the glucosides decreased. Maximum ACN excretion in urine occurred later after BcACN/oatmeal than after BcACN/water administration (3 compared with 2 h). The 4 original ACNs of blackcurrant in their unchanged form, as well as several metabolites, were identified in the urine samples of both groups. The simultaneous intake of food affects ACN absorption and excretion in the urine, but not metabolism.     

COLORIMETRIC CHARACTERIZATION OF EGG YOLK AND EGG YOLK PRODUCTS

Procedures are suggested for COLOR-EYE® evaluation of various yolk forms and products. Chromaticity coordinates derived from COLOR-EYE determinations and the calculated dominant wavelength (λ_d), excitation purity (P_e) and luminous reflectance (Y_CIE) values were also determined for each yolk form and product analyzed. Characteristics of aged yolks included reduced λ_d and P_e and an increase in Y_CIE values; the greatest effects were at the highest levels of pigmentation concentration. Luminous reflectance values were substantially lowered by the addition of sugar and subsequent frozen storage; all other calorimetric criteria were relatively unchanged. Yolk age effects were negligible for the calorimetric characterization of yolks incorporated into product forms. Mayonnaise products provided the most consistent COLOR-EYE readings. These data suggest the feasibility of utilizing the COLOR-EYE, or similar instrumentation, as an objective method for the color characterization of various yolk products. Numerous options are available for converting data into other desired instrumentation values and terminology.     

Modeling the Growth Inhibition Kinetics of Baker's Yeast by Potassium Sorbate Using Statistical Approaches

Potassium sorbate is widely used as a preservative in food products. The inhibition kinetics of yeast growth by potassium sorbate was determined in order to predict the inhibition mechanism and develop an antimicrobial activity model. Linear regression resulted in uncompetitive growth inhibition using different concentrations of nutrients in liquid media containing yeast extract, malt extract (YM broth) and potassium sorbate. At 30°C, constant K_s of YM broth to yeast was 0.254%, maximum growth rate (μ_max) of yeast was 0.008 min^-1, and inhibition constant (K_i) of potassium sorbate to yeast growth was 0.324%. This statistical method could reliably determine the growth inhibition mechanism and kinetics.     

Some characteristics of polyphenol oxidase purified from edible yam (Dioscorea cayenensis-rotundata cv. Longbô) cultivated in Côte d'Ivoire

Polyphenol oxidase (PPO_Y) of edible yam ( Dioscorea cayenensis-rotundata cv. Longbô ) cultivated in Côte d'Ivoire was purified to homogeneity by a procedure that included ion exchange chromatography, ammonium sulphate fractionation, gel filtration and hydrophobic chromatography using dopamine as a substrate. The relative molecular weight of the PPO_Y was estimated to be 94.75 ± 0.63 and 151.56 ± 2.75 kDa by SDS–PAGE and gel filtration on Sephacryl S100 HR, respectively, indicating this PPO_Y as a multimer. Maximal PPO_Y activity occurred at 30 °C and pH 6.6. The enzyme was stable at 25 °C and its pH stability was in the range of 6.0–7.0. The values V _max/ K _m showed that the enzyme had the greatest reactivity towards dopamine among the substrates used. PPO_Y showed no activity toward the monophenols and was completely inhibited by beta-mercaptoethanol, sodium thiosulphate, l -ascorbic acid, sodium bisulphite and l -cysteine. Data generated by this study might help to better prevent edible yam browning.     

Tannins of Grain Sorghum: Luteoforol (Leucoluteolinidin), 3',4,4',5,7-Pentahydroxyflavan

SUMMARY– The principal tannin of sorghum is a leuco-anthocyanin yielding luteolinidin (3',4',5,7-tetrahydroxy-flavylium) when heated with mineral acid. The precursor, luteoforol, has most of the properties of 3',4,4',5,7-penta-hydroxyflavan prepared by reduction of eriodictyol. Luteoforol, when treated with concentrated mineral acid in the cold, gives a purple color with Λ_max550nm. A method for the determination of luteoforol in sorghum, based on this property, is described. The results with a number of varieties of sorghum are compared with those obtained by the AOAC Folin-Denis method. The contribution of luteoforol to the "tannin" so determined varies from 1 to < 25%. Except for one sample of Kaffir corn, which contained leucocyanidin as well as luteoforol, no other tannins were detected. The "tannin" content varied widely, (from 0.05 to 0.67% as tannic acid), a white-skinned variety having the least. The uniformity of commercial samples can be rapidly evaluated by single-grain determinations of luteoforol.     

Influence of vitamin E supplementation on spontaneous oxidized flavour of milk in dairy buffaloes

Twenty-four Murrah buffaloes (60 days pre-partum) were divided into four equal groups (T ₁ , T ₂ , T ₃ and T ₄ ) and were supplemented with 0, 1000, 1500 and 2000 IU α-tocopheryl acetate per day up to 30 days of lactation, and half of these doses from 30 to 60 days of lactation. Milk samples collected fortnightly were analysed for vitamin E, fat, and development of oxidized flavour, with and without copper addition by a panel of judges, and chemically by the thiobarbituric acid test. Scores for oxidized flavour ranged from 0 to 10 with 0–4 as definite, 5–7 as light and 8–10 having no defect. The α-tocopherol content in milk fat (µg/g) averaged 20.55, 25.56, 29.98 and 31.38 in T ₁ , T ₂ , T ₃ and T ₄ groups, respectively. The addition of Cu in the milk significantly increased milk fat oxidation. Better stability of milk in T ₃ and T ₄ groups was observed, which might be due to a higher level of milk α-tocopherol. Addition of 1500 IU α-tocopheryl acetate in the diet of buffaloes helped in improving the oxidative stability of milk.     

MODELING FREQUENCY DISTRIBUTION of STEADY-STATE O2 PARTIAL PRESSURES IN MODIFIED-ATMOSPHERE PACKAGES

A mathematical model was developed to predict the frequency distribution of steady-state O₂ partial pressures in modified-atmosphere packages of fresh and minimally processed fruits and vegetables. Variation in O₂ uptake was predicted to have relatively larger influence on the package O₂ distribution than variation in film permeability to O₂. the model also predicted that the range of O₂ partial pressures in the packages would be larger for the products with lower K_1/2 (a constant in product O₂ uptake equation) values. the percent deviation of package O₂ partial pressures from [O₂]_med increased with [O₂]_med up to about 3 kPa and gradually decreased thereafter. Package O₂ distribution was symmetrical at higher [O₂]_med but became gradually asymmetrical with decreasing [O₂]_med from 6 kPa. the model was used to predict target [O₂]_med for the design of modified-atmosphere packages for different probability levels of having package O₂ being equal to or less than a minimum tolerable level of the product.     

Inhibition of Fungal Growth on Wheat and Rye Bread by Modified Atmosphere Packaging and Active Packaging Using Volatile Mustard Essential Oil

ABSTRACT: Wheat and rye bread artificially inoculated with molds were packed in modified atmospheres of 0%, 50%, 75%, or 100% CO₂ balanced with N₂, and 3 levels of residual O₂, 1%, 0.03%, or <0.01%/O₂-absorber, and stored for 30 to 35 d. Modified atmosphere packaging (MAP) was quantitatively more effective for rye bread because fewer mold species grew at elevated CO₂. However, the major rye bread contaminant, Penicillium roqueforti , was the overall most CO₂-resistant mold and only the use of O₂-absorber could prevent growth of this species. On wheat bread, the most CO₂-tolerant mold was Penicillium commune , growing in 99% CO₂ (with high residual O₂), and Aspergillus flavus was the mold species that grew at lowest O₂ in 75% CO₂ treatment. The spoilage yeast/"chalk mold" Endomyces fibuliger was less affected by the different O₂ levels than the true filamentous molds, and none of the tested MAP treatments could prevent growth, but lag-phase was increased with O₂-absorber on wheat bread and decreased with 1% residual O₂ on rye bread. Experiments with volatile mustard oil showed that A. flavus and Eurotium repens were the most mustard oil-resistant species on wheat and rye bread, respectively. A combination strategy with MAP and mustard oil proved most optimal, and total inhibition was achieved with 2 μL mustard oil/rye bread slice and between 2 and 3 μL/wheat bread. Results indicated that the nature and surface area of the product influences effectiveness of active packaging with mustard oil.