Effect of enzyme‐aided pressing on anthocyanin yield and profiles in bilberry and blackcurrant juices

Bilberries (Vaccinium myrtillus) and blackcurrants (Ribes nigrum) were treated with extensive dosages of commercial cell wall degrading enzyme preparations, i.e. Econase CE, Pectinex Ultra SP‐L, Pectinex Smash, Pectinex BE 3‐L and Biopectinase CCM. The enzymes were dosed based on the polygalacturonase activity. The juice yield was improved in both berries as a result of the enzymatic treatment. The improvement was more pronounced with blackcurrants owing to their thicker cell walls. The impact of the enzymatic treatment on anthocyanins present in the juices was investigated using HPLC‐DAD. The enzyme preparations affected the contents and composition of anthocyanins in the juices. Pectinex Ultra SP‐L, Pectinex Smash, Pectinex BE 3‐L and Biopectinase CCM increased the total content of anthocyanins by 13–41% in the bilberry juices and by 18–29% in the blackcurrant juices. Econase CE, however, produced a dramatic decrease in the total anthocyanin content in the bilberry juice due to its enzyme profile, whereas no such effect was observed with the blackcurrant juice. All the enzyme mixtures tested produced a total or extensive loss of anthocyanidin galactosides in bilberry juice. Commercial enzyme preparations used in the production of berry juices can improve extraction of anthocyanins into the juice. However, they may effectively hydrolyse certain glycosides and thus affect the profile of extracted anthocyanins. Copyright © 2005 Society of Chemical Industry     

Optimising enzymatic maceration in pretreatment of carrot juice concentrate by response surface methodology

In this paper, enzyme preparation for carrot pulp maceration was screened out and enzymatic maceration processing condition of the carrot pulp was optimised by response surface methodology for carrot juice concentrate. Pectinex Smash XXL was the best commercial enzyme preparation than Pectinex Ultra SP‐L, Pectinase FNP‐1 and cellulose FNC‐1 employed in the carrot juice processing in the study. The effect of enzyme concentration and incubation time and their complex interaction on juice β‐carotene content, juice yield and viscosity in the maceration process was studied by using experiments of central composite rotatable design. The results indicated that under the optimal conditions that the enzyme concentration was 100 mL t^?1 and incubation time was 80 min, the juice β‐carotene content was ≥54.2 mg kg^?1, the juice yield ≥63.5% and the juice viscosity ≤2.1 mPa S.     

Protein stability function relations: native β‐lactoglobulin sulphhydryl–disulphide exchange with PDS

Intermolecular sulphhydryl–disulphide exchange with β‐lactoglobulin dimer occurs when this dissociates to form monomers exposing two SH groups. This notion is re‐evaluated in the light of recent structural data suggesting that the degree of SH group exposure in β‐lactoglobulin is unaffected by dissociation. β‐Lactoglobulin was treated with 2,2′‐dipyridyl disulphide (PDS). The rate of sulphhydryl–disulphide exchange was measured at sub‐denaturation temperatures of 25–60 ° C. Parallel studies were conducted by reacting PDS with reduced glutathione (GSH). The SH group of GSH was up to 31 000 times more reactive than β‐lactoglobulin. At pH 7 the reaction activation enthalpy (ΔH^#) and entropy (ΔS^#) was 26 kJ mol⁻¹ and −100 J mol⁻¹ K⁻¹ respectively for GSH. For β‐lactoglobulin, ΔH^# was 157.2 kJ mol⁻¹ and ΔS^# was 254 J mol⁻¹ K⁻¹. At pH 2.6, ΔH^# was 14.4 kJ mol⁻¹ and ΔS^# was −213 J mol⁻¹ K⁻¹ for GSH. The corresponding results for β‐lactoglobulin were 20.3 kJ mol⁻¹ and −147 J mol⁻¹ K⁻¹. These and other thermodynamic results are discussed in terms of the effects of β‐lactoglobulin conformational structure and stability on SH group reactivity. For native β‐lactoglobulin at neutral pH, intermolecular sulphhydryl–disulphide exchange appears to involve the dissociated monomer. SH group activation probably arises from the lower structural stability of the monomer relative to the dimer. At pH 2.6 the mechanism of SH–disulphide exchange does not require protein dissociation and probably involves breathing motions or localised changes in protein structure. © 2000 Society of Chemical Industry     

THERMAL INACTIVATION OF PEA FLOUR LIPOXYGENASE

Lipoxygenase (LOX) was determined in pea flour (9.6% moisture) samples which had been exposed to 90–130°C for 5–45 min, and in crude aqueous extracts (5%; pH 6.7) of previously unheated pea flour which had been exposed to 40–80°C for 5–30 min. The rate of thermal inactivation of pea LOX was shown to follow first-order reaction kinetics. The rate constants, k(min^?1), ranged from 0.005 to 0.252 for the flour, and from 0.068 to 0.267 (60–80°C) for the crude extract. The respective thermodynamic values were: for the energy of activation (E_a), 126.2 and 64.6 kJmole^?1; for the enthalpy of activation (ΔH^?), 123.1 and 61.8 kJmole^?1; for the free energy of activation (Δ^?), 118.0 and 101.4 kJmole^?1. The greater thermostability of LOX in the pea flour under dry-heat conditions, and the observed differences in the E_a, ΔH^? and ΔS^? values, may be accounted for by its possible complexation with other macromolecules and by the structure of the water surrounding the native enzyme.     

A comparative study of polyphenoloxidases from taro (Colocasia esculenta) and potato (Solanum tuberosum var. Romano)

Taro (C. esculenta) is a staple food in many tropical regions. A comparative study of crude polyphenoloxidases from taro (tPPO) and potatoes (pPPO) was carried out to provide information useful for guiding food processing operations. Crude PPO was prepared by cold acetone precipitation using ascorbic acid as antioxidant. The PPO content of taro acetone powder was 770±17 units (mg protein)⁻¹ as compared with 3848±180 units (mg protein)⁻¹ in potato acetone powder. The pH-activity optimum was pH 4.6 for tPPO and pH 6.8 for pPPO. Both enzymes retained >80% activity after incubation at pH 4.5–8 but there was rapid activity loss at pH < 4. The temperature-activity optimum (T_opt) was 30°C for tPPO and 25°C for pPPO with 75 and 27% of their respective maximum activity retained at 60°C. Both tPPO and pPPO were irreversibly inactivated by 10 min heating at 70°C. The activation enthalpy (ΔH^#) and activation entropy (ΔS^#) for tPPO heat-inactivation were 87.4 (±0.1) kJ mol⁻¹ and −56.2 (±4) J mol⁻¹ K⁻¹, respectively. For pPPO, ΔH^# was 59.1 (±0.1) kJ mol⁻¹ whilst ΔS^# was −141 (±4) J mol⁻¹ K⁻¹. The apparent substrate specificity was established from values V_max/K_m as: 4-methylcatechol>chlorogenic acid>dl-dopa>catechol>pyrogallol> dopamine>>caffeic acid for tPPO. There was no detectable activity towards caffeic acid. The substrate specificity for pPPO was: 4-methylcatechol>caffeic acid>pyrogallol>catechol>chlorogenic acid >dl-dopa>dopamine. According to the order of inhibitor effectiveness (sodium metabisulphite>ascorbic acid>NaCl≈ (EDTA), there was a significant lag-phase before increases occurred in the absorbance at 420 nm. Preincubation of PPO with inhibitors increased the extent of inhibition, indicating a direct effect on the structure of the enzyme.     

Thermal Degradation Kinetics of Chlorophyll in Pureed Coriander Leaves

Coriander leaves are widely used in cooking throughout the world. Thermal degradation kinetics of chlorophyll a, b, and total chlorophyll in coriander leaf puree was investigated at varying levels of pH (4.5–8.5) and processing temperature (80–145°C). Coriander puree at pH 4.5 was processed at 80° to 100°C, whereas that at pH 5.5 to 8.5 was processed at 105° to 145°C. Chlorophyll degradation followed first-order reaction kinetics. Good agreement was found between estimated and experimental chlorophyll retention in all cases (R ² > 0.80). Activation energies ranged from 6.57 to 96.00 kJ/mol. Reaction rate and activation energy data indicated that chlorophylls were more stable at alkaline pH. Transition state theory was applied to estimate the enthalpy, entropy, and Gibbs free energy of activation. Enthalpy of activation (ΔH ^#) ranged from 3.46 to 91.99 kJ/mol, whereas entropy of activation (ΔS^#) ranged from −0.265 to −0.047 kJ/(mol K). The overall free energy change was 107.55 kJ/mol. Results indicated that, the compensation effect did not exist for chlorophyll degradation in coriander puree during thermal processing.     

Thermal Chlorophyll Degradation Kinetics of Mint Leaves Puree

Thermal degradation kinetics of chlorophyll ‘a’, ‘b’ and total chlorophyll in mint leaves puree were investigated as function of pH (4.5–8.5) and processing temperature (80–145°C), respectively. Mint puree was processed at 80°to 100°C at pH 4.5, while that at pH 5.5 to 8.5 was processed at 105°to 145°C. Chlorophyll degradation followed the first order reaction kinetics. Good agreement was found between estimated and experimental chlorophyll retention in all cases (R² > 0.86; MRQE < 0.27). Activation energies ranged from 6.45 to 47.67 kJ/mol. Reaction rate and activation energy data indicated that chlorophylls were more stable at alkaline pH. Transition state theory was applied to estimate the enthalpy, entropy and Gibbs free energy of activation. Enthalpy (ΔH^#) ranged from 3.14 to 44.66 kJ/mol, while entropy (ΔS^#) ranged from ?0.157 to ?0.266 kJ/(mol K). The overall free energy change was 105.76 kJ/mol. Results indicated that, the compensation effect did not exist for chlorophyll degradation in mint puree during thermal processing.     

Effect of non‐isothermal processing and moisture content on the anthocyanin degradation and colour kinetics of cherry pomace

Anthocyanins (ACY) and colour changes in cherry pomace under non‐isothermal processing were investigated. Pomace at moisture levels of 70% (MC‐70), 41% (MC‐41) and 25% (MC‐25) was heated at 126.7 °C in a retort for 25, 40 and 60 min. Total ACY, Hunter colour values, total colour difference (ΔE), chroma, hue angle (h°) and browning index (BI) were analysed. Thermal degradation kinetics for colour parameters were determined using zero‐ and first‐order models. ACY degradation increased with heating time and ranged from 34 to 68% for 25 and 60 min heating, respectively. The half‐life of ACY was 38, 33 and 27 min for MC‐70, MC‐41 and MC‐25 pomace, respectively. The ΔE increased with increasing heating time, whereas BI exhibited an inverse trend. Except for ?E for MC‐70, the zero‐order kinetic model showed better fit (R² = 0.85–0.97) to experimental data than the first‐order kinetic model for Hunter colour b values and ?E.     

Kinetic and thermodynamic investigation of mancozeb degradation in tomato homogenate during thermal processing

BACKGROUND: The kinetic and thermodynamic parameters of mancozeb degradation in tomato homogenates under the conditions prevailing in the manufacture of tomato products (at 60–100 °C for 0–60 min) were investigated. A gas chromatography–mass spectrometry method was used to analyse residual mancozeb in tomato homogenate. Ethylenethiourea (ETU), the main toxic degradation product of mancozeb, was measured by high‐performance liquid chromatography (HPLC)–with photodiode array detector (PDA). RESULTS: The degradation of mancozeb and the formation of ETU in tomato homogenates were adequately described as first‐order kinetics. Dependence of the rate constant followed the Arrhenius relationship. Apparent activation energies, temperature coefficients, half time and time to reduce to 90% of the initial value of mancozeb were calculated as kinetic parameters. The thermodynamic parameters of mancozeb were also described as Δg^d = ? 2.440 and 7.074 kJ mol^?1; Δh^d = ? 32.555 and ? 42.767 kJ mol^?1; Δs^d = ? 0.090 and ? 0.150 kJ mol^?1 K^?1; K_e = 0.414 and 9.797 L g^?1 for 333 and 373 K respectively. CONCLUSION: Current findings may shed light on the reduction of mancozeb residue and its toxic degradation product during thermal processing of tomatoes and may also be valuable in awareness and prevention of potential risks from dietary exposure. Copyright © 2011 Society of Chemical Industry     

Pectin lyase is a key enzyme in the maceration of potato tuber

The maceration of potato tuber (Solanum tuberosum cv Bintje) by technical enzyme preparations was examined with the aid of fluorescence microscopy using fluorescein diacetate as a dye. Treatment with Pectinex Ultra-SP-L from Aspergillus aculeatus resulted in a higher release of single viable cells and smaller clumps of cells than treatment with Rohament P from Aspergillus niger. Fractions with high pectin lyase and low polygalacturonase activity obtained by anion exchange chromatography of Pectinex Ultra SP-L showed the highest degree of maceration. When two different cloned polygalacturonases (PG1 and PG2) and a cloned pectin lyase (PL1) from A aculeatus were examined, the highest degree of maceration was observed with PL1. In conclusion, pectin lyase is probably the main enzyme in Pectinex Ultra-SP-L responsible for the maceration of potato tuber. © 1997 SCI.     

Interaction of Protein Isolate with Anthocyanin Extracted from Black Soybean and Its Effect on the Anthocyanin Stability

The interactions between black soybean protein isolate (B‐SPI) and cyanidin 3‐O‐glucoside (C3G), anthocyanin extracted from black soybean coat was investigated under neutral conditions. The fluorescence spectra showed that C3G had fluorescence quenching effects on B‐SPI. Thermodynamic parameters showed that ? G < 0, which demonstrated that the binding was a spontaneous reaction. Since ΔH > 0 and ΔS > 0, the interactions between C3G and B‐SPI was mainly hydrophobic interactions. Fourier infrared spectroscopy results suggested that the contents of α‐helix and β‐sheet structure showed an increasing trend, whereas the β‐angle content displayed a decreasing trend. The degradation of C3G followed first‐order kinetics at 85 °C and 100 °C. After the interactions with B‐SPI, the degradation rate constant was decreased and the half‐life of C3G was prolonged from 70.25 ± 0.90 min to 175.64 ± 38.04 min at 85 °C, from 62.68 ± 1.1 min to 72.51 ± 2.5 min at 100 °C (p < 0.05). The results indicated that the interactions of B‐SPI and C3G improved the thermal stability of C3G under heating conditions.     

Kinetics of flavour and aroma changes in thermally processed cupuaçu (Theobroma grandiflorum) pulp

Changes in ‘fresh’ and ‘cooked‐notes’ during thermal treatment of cupuaçu (Theobroma grandiflorum) pulp were evaluated and modelled. Isothermal experiments in the temperature range of 70–98 °C were carried out and a non‐linear regression was performed to all data to estimate kinetic parameters. ‘Fresh’ and ‘cooked‐notes’ change followed simple first‐order (Ea = 78–82 kJ·mol⁻¹, z = 30–31 °C) and reversible first order (Ea = 80–85 kJ·mol⁻¹) kinetics, respectively. Although ‘cooked‐notes’ were linearly correlated with ‘fresh‐notes’ (R² = 0.99), the former was a better indicator for quality degradation. These results are useful to design pasteurisation processes while minimising sensory changes. © 2000 Society of Chemical Industry     

The Comparative Heat Stability of Bovine β-Lactoglobulin in Buffer and Complex Media

The heat stability of β-lactoglobulin (β-lg) is usually described with reference to a concentration-dependent pseudo-rate constant k. Kinetic and thermodynamic parameters for the irreversible denaturation of β-lg, in a whey protein mixture dissolved in Tris-HCl buffer, were examined over a wide temperature range 75–120°C and for degrees of denaturation [(C_o-C_t)/C_o] up to about 90%. The first-order kinetic model best described β-lg denaturation over the temperature range 75–85°C, whereas the second-order model applies in the range 90–120°C. A comparison between β-lg thermostability in buffer and literature data pertaining to more complex heating media (whey and milk), over the range 75–120°C, was carried out on the basis of changes in activation free energy (ΔG^#), which itself takes into account changes in both activation enthalpy (ΔH^#) and activation entropy (ΔS^#). It was found that the thermal stability of β-lg in different media, and irrespective of the kinetic model assumed in the present study, can be ranked: buffer ≪ whey>milk for the temperature range 75–85°C. On the contrary, at the higher temperature range, 90–120°C, the ranking is buffer<whey<milk, when using the second-order model to describe the present data. For such comparisons to be valid the initial concentration of β-lg in various studies, as well as the reaction order applied, should be taken into consideration. © 1997 SCI.     

Patulin Adsorption Kinetics on Activated Carbon, Activation Energy and Heat of Adsorption

The kinetics of adsorption of patulin on activated carbon were studied at different initial patulin concentrations (100–400 ppb) for the temperature range 20–80°C. Apparent adsorption rate constants (k_aapp) were changed from 1.07 × 10^?3 to 1.86 × 10^?3 g^?1 min^?1 while the temperature increased from 20 to 80°C. For equilibrium adsorption curves; the Langmuir model was attempted and model parameters (K and Q°) were obtained for different temperatures. Energy of activation and heat of adsorption were determined in a batch adsorption system (E_a= 2.02 kcal/mol and ΔH = 2.24 kcal/mol). The adsorption occurred endothermically and by physical mechanisms.     

Degradation Kinetics of Anthocyanin in Blueberry Juice during Thermal Treatment

ABSTRACT: The kinetics of anthocyanin degradation in blueberry juice during thermal treatment at 40, 50, 60, 70, and 80 °C were investigated in the present study. Anthocyanin degradation was analyzed up to the level of 50% retention using a pH differential method. The degradation of anthocyanin at each temperature level followed a first-order kinetic model, and the values of half-life time (t_1/2) at temperatures of 40, 50, 60, 70, and 80 °C were found to be 180.5, 42.3, 25.3, 8.6, and 5.1 h, respectively. The activation energy value of the degradation of the 8.9 ° Brix blueberry juice during heating was 80.4 kJ·mol⁻¹. The thermodynamic functions of activation (ΔG, ΔH, and ΔS) have been determined as central to understanding blueberry degradation.     

Inactivation of bacteriophages by thermal and high-pressure treatment

Dairy companies commonly experience fermentation failures due to bacteriophages that are spread mainly by milk, whey or air. Heat or high-pressure treatment may potentially reduce the phage titre, but further knowledge about the inactivation kinetics is desirable. Inactivation experiments were carried out with the commonly occurring lactococcal phages P001 and P008. Phage suspensions in calcium-enriched M17-broth were heated at 55–80 °C, or high-pressure treated at up to 600 MPa. Kinetic analysis showed that the order of inactivation reaction was above 1; thus, inactivation kinetics were approximated by a non-linear regression model. The Arrhenius parameters, rate constant, k_p,T, and activation energy, E_A (for heat treatments), and the volume of activation, ΔV^# (for pressure treatments) were calculated. Both measured and calculated results indicate that phage P008 was the more heat- and pressure-resistant of the two. By combining the results from heat and pressure inactivations, a pressure–temperature diagram for phage P008 was established.     

The utility of a fungal bibonuclease for reducing the nucleic acid content of permeabilized yeast cells

The main drawback of yeast biomass as a source of protein for human consumption is its high nucleic acid content. The present study deals with the development of a process for reducing the nucleic acid content of strains of Saccharomyces cerevisiae, Candida utilis, C.tropicalis and C.lipolytica by treating with an RNase of Aspergillus candidus strain Ml6a. The cells were permeabilised either by heat‐treatment at 95°C for 5 min. or by treatment with chloroform for 6h followed by a heat treatment at 65°C for 3 min. The former pretreatment was sufficient for C. utilis and C.tropicalis strains whereas S.cerevisiae required the latter treatment. The optimum conditions for the enzymatic treatment were a pH of 4.5–5.0, temperature of 45–55°C, incubation period of 60–90 min and an enzyme to cell ratio of 1:6,000 (w/v). Crude enzyme preparations showed a better activity than the pure enzyme. Under optimal conditions 80–85% of the total nucleic acid could be removed from yeast cells by the enzymatic treatment without any significant concomitant loss of protein.     

Study of pulsed light inactivation and growth dynamics during storage of Escherichia coli ATCC 35218, Listeria innocua ATCC 33090, Salmonella Enteritidis MA44 and Saccharomyces cerevisiae KE162 and native flora in apple,orange and strawberry juices

The response of some inoculated strains and native flora to PL treatment (Xenon lamp, 3 pulses s^?1, 10 cm distance from the lamp, 71.6 J cm^?2) in apple, orange and strawberry fresh juices with different absorbance, turbidity and particle size was investigated. Microbial growth dynamics during 12‐day storage (5 °C) of PL‐treated juices was also evaluated. PL treatments provoked 0.3–2.6 log reductions for inoculated microorganisms and 0.1–0.7 for native flora. High turbidity and particles with high UV absorbance seemed to play a major role in the PL efficiency compared to particle size. Cold storage of PL‐processed juices provoked an increase in Salmonella Enteritidis and Listeria innocua inactivation, achieving 5.0–8.0 log reductions, while no recovery of Escherichia coli and retardation for yeast growth was observed, compared to untreated samples. This study gives valuable information regarding the influence of juice variables on PL effectiveness and emphasises the beneficial effect of a postcold storage on microbial safety of PL‐treated juices.     

Purification and immobilisation of inulinase from Aspergillus candidus for producing fructose

A high‐inulinase‐producing strain of Aspergillus candidus (10 units cm⁻³ of medium) for producing fructose from inulin has been identified. The extracellular inulinase from this fungi was purified 56‐fold by ammonium sulphate fractionation, DEAE cellulose and Sephadex G‐150 column chromatography. Invertase to inulinase ratio of 1.8 in culture filtrate was reduced to 0.14 in the purified preparation. The pH and temperature optima were 5.5 and 45°C, respectively. The molecular weight of inulinase was determined as 54±4 kDa. K_m of inulinase with inulin as a substrate was 3.8 mmole dm ⁻³. The purified preparation produced only fructose as the product of inulin, indicating that inulinase has primarily exo‐inulinase activity. Inulinase was immobilised on chitin and casein using glutaraldehyde as a linking agent and on cellulose using FeCl₃‐HCl as a metal chelation agent. Maximum immobilisation of 45.8% was achieved on cellulose. All three immobilised preparations had a higher temperature optima of 55°C. The inulinases immobilised on cellulose and casein were stable at pH 5–7. The cellulose‐immobilised preparation was more stable than the other two preparations after heating for 1 h at 55°C. © 1999 Society of Chemical Industry     

Validation of immunodetection (ELISA) of ricin using a biological activity assay

Abstract: The suitability of enzyme‐linked immunosorbent assay (ELISA) for residual ricin toxicity determination was investigated in this study. Ricin was thermally treated at 80 to 90 °C for up to 9 min, and its residual concentration was determined by means of a commercial ELISA kit, and its bioactivity (amount of adenine released from DNA) was determined by means of a biological activity assay (BAA). Results showed that inactivation of ricin followed 1st‐order kinetics. The half‐life values for loss of bioactivity at 80, 85, and 90 °C were 1.93, 0.65, and 0.41 min, respectively. Similarly, the half‐life values for reduction in ricin concentration determined by ELISA were 3.06, 0.79, and 0.43 min, respectively. The half‐lives determined by both assays were only significantly different at 80 °C. The Z, Q₁₀, and Arrhenius activation energy values determined by both assays were dissimilar: 11.74 ?C, 7.12 and 50.1 kcal/mol, respectively, by ELISA; and 14.87 °C, 4.71 and 39.5 kcal/mol, respectively, by BAA. Nevertheless, our findings indicate that the 2 assays were highly correlated (R²= 1), and it can be concluded that ELISA would be a reliable method for detecting residual toxicity of heat‐treated ricin based on fraction lost. Practical Application: The results of this study indicate that immunodetection, even though not a direct measurement of the biological activity of ricin, is suitable for determining the residual bioactivity of ricin since immunodetection and the biological activity assay used in this investigation were highly correlated. Therefore, ELISA can be used for routine assessment of residual activity or toxicity of ricin in thermally treated foods.