A kinetic study on the thermal inactivation of barley malt α-amylase and ß-amylase during the mashing process.

To obtain an efficient conversion of starch into fermentable sugars and dextrins during the brewing process, mashing time-temperature profiles need to promote starch gelatinisation and enzyme activity while avoiding thermal inactivation of the amylases. This study focused on the second part of this balance by investigating the thermal stability of α-amylase and ß-amylase of Planet barley malt throughout mashing. Thermal inactivation in wort was modelled for both enzymes resulting in the estimation of thermal inactivation kinetic parameters such as rate constant of thermal inactivation kT (the rate of thermal inactivation of an enzyme at a constant temperature), activation energy for thermal inactivation Ea, decimal reduction time DT (the time needed to inactivate 90% of the enzyme activity at a given temperature) and the z-value. First-order inactivation was observed for α-amylase. For ß-amylase, fractional conversion inactivation occurred with a residual fraction of 13% of the ß-amylase activity that remained after prolonged heating at 72.5 °C. The ß-amylase protein population hence seems to consist of thermolabile and thermostable isoforms. The kinetic parameters for thermal inactivation of the enzymes were used to predict their residual activities throughout a laboratory-scale mashing process. The predicted residual activities met the experimentally determined residual enzyme activities closely, except for ß-amylase at temperatures higher than 72.5 °C. The results obtained in this work allow designing new mashing processes or tailoring existing processes towards variability in the input material, barley malt, without the need for trial-and-error experiments.

Digestion kinetics of lipids and proteins in plant-based shakes: Impact of processing conditions and resulting structural properties.

In this work, plant-based shakes were prepared (5% oil, 6% protein, 1% lecithin, 88% water) (w/w) using two processing techniques (i) only mixing versus (ii) mixing followed by high pressure homogenisation, as well as two processing sequences (i) adding all ingredients together versus (ii) stepwise addition of ingredients. Shakes only mixed consisted of large, irregular particles (1-100 µm). Eventually, this resulted in a relatively low lipid and protein digestion extent after 2 h of gastric pre-digestion (9% and < 1%, respectively). In contrast, shakes that were subjected to high pressure homogenisation displayed small, homogeneous particles (<10 µm). Besides, lipids and proteins were digested to a high extent in the stomach (40% and 10%, respectively). The small intestinal digestion kinetics indicated a significant impact of proteins on lipid digestion kineticsbutno significant effect of lipids on protein digestion kinetics. The results highlighted the relevance of food processing on macronutrient (micro)structure and further gastrointestinal functionality.

Structural and emulsion stabilizing properties of pectin rich extracts obtained from different botanical sources.

The presented research studied the emulsifying and emulsion stabilizing capacity of pectin samples isolated from different plant origin: apple, carrot, onion and tomato. The acid extracted pectin samples showed distinct structural properties. Specifically, apple pectin showed a high degree of methylesterification (78.41 ± 0.83%), carrot pectin had the lowest concentration of other co-eluted cell wall polymers, onion pectin displayed a bimodal molar mass distribution suggesting two polymer fractions with different molar mass and tomato pectin was characterized by a high protein content (16.48 ± 0.05%). The evaluation of the emulsifying and emulsion stabilizing potential of the pectin samples included investigating their ability to lower the interfacial tension next to a storage stability study of pectin stabilized o/w emulsions. Creaming behavior as well as the evolution of the oil droplet size were thoroughly examined during storage using multiple analytical techniques. Overall, smaller oil droplet sizes were obtained at pH 2.5 compared to pH 6.0 indicating better emulsifying capacity at lower pH. The lowest emulsion stability was observed in emulsions formulated with tomato pectin in which weak flocculation and relatively fast creaming affected emulsion stability. Onion pectin clearly showed the most promising emulsifying and emulsion stabilizing potential. At both pH conditions, emulsions stabilized by the onion pectin sample displayed highly stable oil droplet sizes during the whole storage period. The presence of the two polymer fractions in this sample can play an important role in the observed stability. In future work, it could be evaluated if both fractions contribute to emulsion stability in a synergistic way. In conclusion, this work showed that pectin samples extracted from different plant origin display diverse structural properties resulting in varying emulsifying and emulsion stabilizing potential. Polymer molar mass potentially plays a major role in the structure-function relation.

Starch hydrolysis during mashing: A study of the activity and thermal inactivation kinetics of barley malt α-amylase and ß-amylase.

Hydrolysis of starch is key in several industrial processes, including brewing. Here, the activity and inactivation kinetics of amylases throughout barley malt mashing are investigated, as a prerequisite for rational optimisation of this process. Varietal differences were observed in the activity of α- and ß-amylases as a function of temperature for six barley and malt varieties. These differences were not reflected in the resulting wort composition after mashing, using three isothermal phases of 30 min at 45 °C, 62 °C and 72 °C with intermediate heating by 1 °C/min. Thermal inactivation kinetics parameters determined for α- and ß-amylases of an industrially relevant malt variety in a diluted system showed that enzymes were inactivated at lower temperatures than expected. The obtained kinetic parameters could predict α-amylase, but not ß-amylase inactivation in real mashing conditions, suggesting that ß-amylase stability is enhanced during mashing by components present or formed in the mash.

Pectin influences the kinetics of in vitro lipid digestion in oil-in-water emulsions.

Oil-in-water emulsions were prepared with 5% (w/v) carrot-enriched olive oil and stabilized with Tween 80 (TW), phosphatidylcholine (PC), citrus pectin (CP) or a combination of these emulsifiers. Additionally, the methylesterification degree (DM) of citrus pectin was modified, resulting in three different studied pectin structures: CP82, CP38 and CP10. All initial emulsions presented small initial oil droplet sizes and were submitted to an in vitro simulated gastric and small intestinal phase. The latter was executed in a kinetic way to determine the time dependency of the lipolysis reaction, micelle formation and carotenoid bioaccessibility. The results showed that the pectin DM mainly influenced the reaction rate constants, while the emulsifier (combination) determined the extent of lipolysis and carotenoid bioaccessibility. Moreover, a direct relation was observed between the lipolysis reaction and bioaccessibility extent. The presented study showed that targeted emulsion design can be used to tailor lipid digestion kinetics.

Emulsion stability during gastrointestinal conditions effects lipid digestion kinetics.

Oil-in-water emulsions were prepared with carrot- or tomato-enriched olive oil (5%w/v) and stabilized with Tween80 or sucrose esters (0.5%w/v) with different hydrophilic-lipophilic balance (8; 11 or 16). All emulsions had similar initial oil droplet sizes and were submitted to simulated gastrointestinal conditions using a kinetic digestion procedure. Sucrose esters induced an unstable system after gastric conditions leading to coalesced oil droplets, while Tween80 emulsions remained stable. Emulsion particle sizes at the end of the gastric phase were directly associated with the lipolysis kinetics during the intestinal phase. Moreover, a direct relationship was observed between lipolysis and carotenoid micellarisation for all emulsions, and depended mainly on the surfactant structure used. Tween80 emulsions led to a higher lipolysis extent (53-57%) and carotenoid bioaccessibility (17-42%) compared to sucrose ester emulsions (33-52% and 9-27%, respectively). These findings show the importance of the emulsifier structure and emulsion stability during gastrointestinal conditions in modulating lipolysis kinetics.

Lipid digestion, micelle formation and carotenoid bioaccessibility kinetics: Influence of emulsion droplet size.

Carotenoid-enriched oil-in-water emulsions with different droplet sizes (small: d43 0.72µm; medium: d43 1.9µm; large: d43 15.1µm) were subjected to simulated gastrointestinal conditions. The kinetics of lipolysis, micelle formation and carotenoid bioaccessibility were monitored during the intestinal phase. The rates of all three processes increased with decreasing droplet size. The large droplet size emulsion contained undigested oil at the end of digestion, whereas an almost complete hydrolysis was observed for the other two emulsions. The sub-micron emulsion presented a higher conversion of MAGs to FFAs during digestion, which led to a higher concentration of FFAs in the mixed micelles. The incorporation of carotenoids into mixed micelles occurred faster and reached a higher final value for the small droplet size emulsion, leading to final carotenoids bioaccessibility values of around 70%. This work provides valuable information for developing in silico models to simulate the lipid digestibility and carotenoid bioaccessibility.

L-ascorbic acid improves the serum folate response to an oral dose of [6S]-5-methyltetrahydrofolic acid in healthy men.

OBJECTIVE: To investigate the effect of simultaneous administration of [6S]-5-methyltetrahydrofolic acid ([6S]-5-CH(3)H(4)PteGlu) with L-ascorbic acid (L-AA) on serum folate concentrations in healthy male subjects. SUBJECTS AND METHODS: A total of nine healthy male volunteers were recruited. Serum folate concentrations were measured before and up to 8 h after administration of each treatment (1) placebo, (2) 343 microg [6S]-5-CH(3)H(4)PteGlu), (3) 343 microg [6S]-5-CH(3)H(4)PteGlu) with 289.4 mg L-AA and (4) 343 microg [6S]-5-CH(3)H(4)PteGlu) with 973.8 mg L-AA (n=10 samples per treatment). RESULTS: Serum folate concentrations significantly increased compared with baseline values, starting from 30 min after [6S]-5-CH(3)H(4)PteGlu administration and remained significantly higher than baseline values during the first 6 h for treatments 3 and 4, and during the first 4 h for treatment 2. Maximal serum folate responses were observed between 0.5 and 1.5 h after [6S]-5-CH(3)H(4)PteGlu consumption and significantly differed between treatments 2 and 4 (P<0.05). When [6S]-5-CH(3)H(4)PteGlu was concurrently administered with 289.4 or 973.8 mg L-AA, the total serum folate response, calculated as the area under the curve (AUC), was significantly improved (46.5+/-4.0 and 53.0+/-4.0 vs 34.3+/-3.8 h nmol/l, P<0.05). No significant difference in AUC was found between the 289.4 and the 973.8 mg L-AA treatments. CONCLUSIONS: Administration of a physiological dose of [6S]-5-CH(3)H(4)PteGlu with L-AA significantly improved the measured serum folate response in folate saturated healthy men.

Inactivation kinetics of alkaline phosphatase and lactoperoxidase, and denaturation kinetics of beta-lactoglobulin in raw milk under isothermal and dynamic temperature conditions.

A detailed kinetic study of alkaline phosphatase, lactoperoxidase and beta-lactoglobulin was carried out in the context of identifying intrinsic time-temperature indicators for controlling the heat processing of milk. The heat inactivation or denaturation of alkaline phosphatase, lactoperoxidase and beta-lactoglobulin under isothermal conditions was found to follow first order kinetics. Experimental results were analysed using both a two step linear regression and a one step non-linear regression method. Results obtained using the two statistical techniques were comparable, but the 95% confidence interval for the predicted values was smaller when the one step non-linear regression method was used, indicating its superiority for estimating kinetic parameters. Thermal inactivation of alkaline phosphatase and lactoperoxidase was characterized by z values of 5.3 deg C (D60 degrees C = 24.6 min) and 4.3 deg C (D71 degrees C = 38.6 min) respectively. For the denaturation of beta-lactoglobulin we found z values of 7.9 deg C (D7.5 degrees C = 49.9 min) in the temperature range 70-80 degrees C and 24.2 deg C (D85 degrees C = 3.53 min) in the range 83-95 degrees C. Dref and z were evaluated under dynamic temperature conditions. To estimate the statistical accuracy of the parameters, 90% joint confidence regions were constructed.

Modeling conductive heat transfer during high-pressure thawing processes: determination of latent heat as a function of pressure.

A numerical heat transfer model for predicting product temperature profiles during high-pressure thawing processes was recently proposed by the authors. In the present work, the predictive capacity of the model was considerably improved by taking into account the pressure dependence of the latent heat of the product that was used (Tylose). The effect of pressure on the latent heat of Tylose was experimentally determined by a series of freezing experiments conducted at different pressure levels. By combining a numerical heat transfer model for freezing processes with a least sum of squares optimization procedure, the corresponding latent heat at each pressure level was estimated, and the obtained pressure relation was incorporated in the original high-pressure thawing model. Excellent agreement with the experimental temperature profiles for both high-pressure freezing and thawing was observed.

Lipoxygenase inactivation in green beans (Phaseolus vulgaris L.) due to high pressure treatment at subzero and elevated temperatures.

The kinetics of lipoxygenase (LOX) inactivation in green beans due to high-pressure treatment were studied in the pressure-temperature area of 0.1 up to 650 MPa and -10 up to 70 degrees C for systems with different levels of food complexity, i.e., in green bean juice and intact green beans (in situ study). For both systems, LOX was irreversibly inactivated by high-pressure treatment combined with subzero and elevated temperatures and the inactivation could be described as a first-order reaction. At ambient pressure, in situ LOX was less thermostable than in the juice at temperatures below 68 degrees C whereas the stability ranking was reverse at temperatures above 68 degrees C. At temperatures below 63 degrees C, sensitivity of the inactivation rate constants to temperature changes was on the same order of magnitude in the juice and in situ, while at higher temperature it was lower in situ. The pressure needed to obtain the same rate of LOX inactivation at a given temperature was lower in situ than in the juice. Application of high-pressure treatment at low/subzero temperature resulted in an antagonistic effect on LOX inactivation for both systems, whereas no such effect was found above room temperature. The pressure-temperature dependence of the LOX inactivation rate constants in green beans was successfully modeled.

Inactivation of orange pectinesterase by combined high-pressure and -temperature treatments: a kinetic study.

Pressure and/or temperature inactivation of orange pectinesterase (PE) was investigated. Thermal inactivation showed a biphasic behavior, indicating the presence of labile and stable fractions of the enzyme. In a first part, the inactivation of the labile fraction was studied in detail. The combined pressure-temperature inactivation of the labile fraction was studied in the pressure range 0.1-900 MPa combined with temperatures from 15 to 65 degrees C. Inactivation in the pressure-temperature domain specified could be accurately described by a first-order fractional conversion model, estimating the inactivation rate constant of the labile fraction and the remaining activity of the stable fraction. Pressure and temperature dependence of the inactivation rate constants of the labile fraction was quantified using the Eyring and Arrhenius relations, respectively. By replacing in the latter equation the pressure-dependent parameters (E(a), k(ref)(T)()) by mathematical expressions, a global model was formulated. This mathematical model could accurately predict the inactivation rate constant of the labile fraction of orange PE as a function of pressure and temperature. In a second part, the stable fraction was studied in more detail. The stable fraction inactivated at temperatures exceeding 75 degrees C. Acidification (pH 3.7) enhanced thermal inactivation of the stable fraction, whereas addition of Ca(2+) ions (1 M) suppressed inactivation. At elevated pressure (up to 900 MPa), an antagonistic effect of pressure and temperature on the inactivation of the stable fraction was observed. The antagonistic effect was more pronounced in the presence of a 1 M CaCl(2) solution as compared to the inactivation in water, whereas it was less pronounced for the inactivation in acid medium.

Effect of temperature and/or pressure on tomato pectinesterase activity.

The activity of tomato pectinesterase (PE) was studied as a function of pressure (0.1-900 MPa) and temperature (20-75 degrees C). Tomato PE was rather heat labile at atmospheric pressure (inactivation in the temperature domain 57-65 degrees C), but it was very pressure resistant. Even at 900 MPa and 60 degrees C the inactivation was slower as compared to the same treatment at atmospheric pressure. At atmospheric pressure, optimal catalytic activity of PE was found at neutral pH and a temperature of 55 degrees C. Increasing pressure up to 300 MPa increased the enzyme activity as compared to atmospheric pressure. A maximal enzyme activity was found at 100-200 MPa combined with a temperature of 60-65 degrees C. The presence of Ca(2+) ions (60 mM) decreased the enzyme activity at atmospheric pressure in the temperature range 45-60 degrees C but increased enzyme activity at elevated pressure (up to 300 MPa). Maximal enzyme activity in the presence of Ca(2+) ions was noted at 200-300 MPa in combination with a temperature of 65-70 degrees C.

Modeling conductive heat transfer and process uniformity during batch high-pressure processing of foods.

A numerical model for predicting conductive heat transfer during batch high hydrostatic pressure (HHP) processing of foods was developed and tested for a food simulator (agar gel). For a comprehensive evaluation of the proposed method, both "conventional" HHP processes, HHP processes with gradual, step-by-step pressure buildup and pressure release, and pressure cycling HHP processes were included. In all cases, good agreement between experimental and predicted temperature profiles was observed. The model provides a very useful tool to evaluate batch HHP processes in terms of uniformity of any heat- and/or pressure-related effect. This is illustrated for inactivation of Bacillus subtilis alpha-amylase, an enzymatic model system with known pressure-temperature degradation kinetics.

Kinetics of pressure inactivation at subzero and elevated temperature of lipoxygenase in crude green bean (Phaseolus vulgaris L.) extract.

Lipoxygenase (LOX) in crude green bean extract was irreversibly inactivated by pressure treatments combined with subzero or elevated temperature. LOX inactivation was described accurately assuming a first-order reaction. In the entire pressure-temperature domain studied (200 to 700 MPa and -10 to 60 degrees C), an increase in pressure at constant temperature enhanced the LOX inactivation rate, whereas at constant pressure, an increase in reaction rate was obtained by either increasing or decreasing temperature at 20 degrees C. At elevated pressure, LOX exhibited the greatest stability around 20 degrees C. Also the pressure dependence of the inactivation rate constants for LOX was the highest around 20 degrees C. On the basis of the estimated LOX inactivation rate constants, an iso-rate contour diagram as a function of pressure and temperature was constructed, and an empirical mathematical model describing the combined pressure-temperature dependence of the LOX inactivation rate constants was formulated.

Thermal and combined pressure--temperature inactivation of orange pectinesterase: influence of pH and additives.

Inactivation of commercially available orange pectinesterase (PE) was investigated under isothermal and isothermal-isobaric conditions. In both cases, inactivation data could be accurately described by a fractional conversion model. The influence of enzyme concentration, pH, Ca(2+) concentration, and sucrose on the inactivation kinetics was studied. Enzyme stability against heat and pressure increased by increasing enzyme concentration. An increased Ca(2+) concentration caused sensitization to temperature and increased the residual fraction active PE after thermal treatment. To the contrary, in the case of pressure treatment, decreasing Ca(2+) concentrations increased pressure inactivation. The remaining fraction active PE after pressure treatment was not influenced by the addition of Ca(2+) ions. Acidification accelerated thermal as well as pressure-temperature inactivation, whereas in the presence of sucrose an increased temperature and pressure stability of orange PE was observed. Sucrose had no influence on the remaining activity after thermal treatment, but it increased the residual fraction after pressure treatment. The remaining fraction was for all additives studied independent of the pressure and temperature level applied except for the inactivation in an acid medium, when a decrease of the residual fraction was observed with increasing temperature and pressure.

Single, combined, or sequential action of pressure and temperature on lipoxygenase in green beans (Phaseolus vulgaris L.): a kinetic inactivation study.

The objective of this investigation was to study kinetically the effect of pressure and temperature either as a single, as a combined, or as a sequential action on lipoxygenase (LOX) inactivation in crude green beans extract. The LOX isozymes in green beans extract had a different heat sensitivity but a similar pressure stability: two fractions following apparent first-order reactions, i.e., a heat-labile fraction and a heat-stable fraction, were observed in studies on its thermostability, whereas only one fraction following a first-order reaction was noticed in studies on its pressure stability. At ambient pressure, irreversible LOX inactivation was studied in a temperature range from 55 to 70 degrees C. At room temperature, pressures around 500 MPa were required in order to inactivate LOX in green beans extract. The effect of a pressure or a thermal pretreatment on LOX thermo- or barostability, respectively, was also investigated but no significant differences in inactivation kinetics due to the pretreatment were observed.

Kinetics of chlorophyll degradation and color loss in heated broccoli juice.

Degradation of chlorophyll in broccoli juice occurred at temperatures exceeding 60 degrees C. Chemical analysis revealed that degradation of chlorophyll a and b to pheophytin a and b, respectively, followed first-order kinetics and that chlorophyll a was more heat sensitive than chlorophyll b. Temperature dependencies of chlorophyll a and b degradation rate constants could be described by Arrhenius equations with activation energies (E(a)) of 71.04 +/- 4.89 and 67.11 +/- 6.82 kJ/mol, respectively. Objective greenness measurements, using the -a value as the physical property, together with a fractional conversion kinetic analysis, indicated that green color degradation followed a two-step process. Kinetic parameters for the first degradation step were in accordance with the kinetic parameters for pheophytinization of the total chlorophyll content, as determined by chemical analysis (E(a) approximately 69 kJ/mol). The second degradation step, that is, the subsequent decomposition of pheophytins, was characterized by an activation energy of 105.49 +/- 4.74 kJ/mol.

Soybean lipoxygenase inactivation by pressure at subzero and elevated temperatures.

Soybean lipoxygenase (LOX) inactivation [0.4 mg/mL in Tris-HCl buffer (0.01 M, pH 9)] was studied quantitatively under constant pressure (up to 650 MPa) and temperature (-15 to 68 degrees C) conditions and kinetically characterized by rate constants, activation energies, and activation volumes. The irreversible LOX inactivation followed a first-order reaction at all pressure-temperature combinations tested. In the entire pressure-temperature area studied, LOX inactivation rate constants increased with increasing pressure at constant temperature. On the contrary, at constant pressure, the inactivation rate constants showed a minimum around 30 degrees C and could be increased by either a temperature increase or decrease. On the basis of the calculated rate constants at 102 pressure-temperature combinations, an iso-rate contour diagram was constructed as a function of pressure and temperature. The pressure-temperature dependence of the LOX inactivation rate constants was described successfully using a modified kinetic model of Hawley.