Inactivation Of Pectin Methylesterase Research Articles

Effect of calcium and sucrose on the thermal inactivation of pectin methylesterase from tomato fruits: A kinetic study. [pdf

Effect of calcium and sucrose on the thermal inactivation of pectin methylesterase from tomato fruits: A kinetic study. [pdf

Pulsed electric field processing of fruit juices with inactivation of enzymes with new inactivation kinetic model and determination of changes in quality parameters

Pulsed electric fields (PEF) processing of fresh grape, orange and tomato juices by 17.2–24.1 kV/cm electric field strengths, 130.0–1040.5 μs treatment times and 1.43–29.13 J/kg energies with respect to determination of the changes in the physical and bioactive properties, color parameters in addition to inactivation of pectin methyl esterase (PME), polyphenol oxidase (PPO), and lipoxygenase (LOX) enzymes were studied. In general, decrease in pH, increase in conductivity (except for grape juice), total phenolic substance content, and total antioxidant capacity with no significant change (p > 0.05) in total soluble solids were observed by PEF treatment for all juices. Except for hue value for orange juice, no significant changes (p > 0.05) were observed in color values. Inactivation of the enzymes were increased with increased energy, and calculated electrical activation energies for PME, PPO, and LOX enzymes were 8.31, 7.06, and 4.31 kJ/mol, respectively. Industrial relevanceEnzymes in addition microorganisms causing quality degradation is of great concern in fruit juice industry as most of the enzymes are resistant to processing technologies. Increased PEF treatment time provided over 95% inactivation on PME, PPO, and LOX enzymes with no adverse effect on quality properties of grape, orange, and tomato juices revealing a viable alternative to juice processing industry.

Inactivation mechanisms on pectin methylesterase by high pressure processing combined with its recombinant inhibitor

High pressure processing (HPP) juice often experiences cloud loss during storage, caused by the activity of pectin methylesterase (PME). The combination of HPP with natural pectin methylesterase inhibitor (PMEI) could improve juice stability. However, extracting natural PMEI is challenging. Gene recombination technology offers a solution by efficiently expressing recombinant PMEI from Escherichia coli and Pichia pastoris. Experimental and molecular dynamics simulation were conducted to investigate changes in activity, structure, and interaction of PME and recombinant PMEI during HPP. The results showed PME retained high residual activity, while PMEI demonstrated superior pressure resistance. Under HPP, PMEI's structure remained stable, while the N-terminus of PME's α-helix became unstable. Additionally, the helix at the junction with the PME/PMEI complex changed, thereby affecting its binding. Furthermore, PMEI competed with pectin for active sites on PME, elucidating. The potential mechanism of PME inactivation through the synergistic effects of HPP and PMEI.

Modeling and experimental validation of the time-temperature profile, pectin methylesterase inactivation, and ascorbic acid degradation during the continuous flow microwave-assisted pasteurization of orange juice

This work evaluated the effect of microwave-assisted pasteurization on pectin methylesterase (PME) and ascorbic acid (AA) in orange juice (OJ) and proposed a model to predict the time-temperature profile of the process and consequent effect on inactivation and quality. Fresh OJ was pasteurized in a pilot-scale unit at four processing temperatures (70, 80, 90 and 100) °C, five holding times, using two heating systems (conventional and microwave). The experimental overall heat transfer coefficient was determined from the heat loads and correlated with Reynolds and Prandtl numbers. The best-adjusted correlation was used to predict the time-temperature profile considering a unidimensional mathematical model for flow and heat transfer. Predicted and experimental temperatures showed differences below 0.9 °C for all conditions. The model was able to predict PME inactivation and AA retention, indicating that microwave pasteurization of OJ can be as efficient as conventional heating on PME inactivation with less impact on AA.

Evaluation of the microwave-assisted enzymatic inactivation process of tomato pulp

In this study, the thermal and non-thermal effects of microwaves on the enzymatic inactivation of pectin methylesterase in the tomato pulp were validated. An amount of 30 kg of tomato pulp was subject to two different types of treatments: thermal effects and then non-thermal effects using microwaves, implementing a system that allowed simultaneous cooling of microwave heating. An experimental rotational central composite design was used to analyze the effect of the interactions of the variables involved in the process. The activity analysis of the PME enzyme, pH, titratable acidity, total soluble solid, and color was performed on both processed and non-processed tomato pulps, which showed that the heating process with thermal effect did not significantly change physical-chemical properties and were found to be more effective, reaching the enzymatic inactivation of 99% and the temperature of 80ºC after 425 s of processing. However, the non-thermal analysis with temperatures at a maximum of 40ºC attained 56.8% of enzymatic inactivation.

Effect of hydrodynamic cavitation processing on orange juice physicochemical and nutritional properties

Effect of hydrodynamic cavitation (HC) processing on pH, °brix, separation index, viscosity and color properties, retention of nutritional bioactive compounds (Vitamin-C, total phenolics, and antioxidant), and inactivation of enzyme pectin methyl esterase (PME) in freshly expressed juice from Brazilian oranges was investigated. HC at 4 bar pressures with a treatment time of 0, 10, 20, 30, 40, 50, 60, and 90 min at low temperatures (42 °C) were studied. There was no significant change in the values of pH, and TSS was noted in HC-treated juices. Further decreased flow behavior index while increased values of flow consistency index were recorded for the juice treated with HC showing a significant physical modification of the juice. Degradation and loss in vitamin C and total phenolics were observed when juices were HC-treated for a longer time. Therefore, it was concluded that HC could be used as a non-thermal method for improving the stability and physicochemical properties of Brazilian orange juice.

Application of thermosonication for guava juice processing: Impacts on bioactive, microbial, enzymatic and quality attributes

The present work explores different conditions of thermosonication (TS) processing that would ensure microbiological and enzymatic safety for guava juice while simultaneously maximizing the preservation of its quality attributes. The guava juice was subjected to TS treatment (frequency: 40 kHz; power: 200 W; Temperature: 40, 60, and 80 °C; Time: 2, 6 and 10 min) and was compared with fresh and pasteurized (90 °C/60 s) juice samples. The objectives of the research work were to determine the effect of thermosonication on the quality attributes such as total soluble solids (TSS), pH, titratable acidity, cloud value, color attributes, total phenolic contents, total flavonoid contents, antioxidant activity, ascorbic acid levels, enzymatic, microbiological, and sensory properties. The thermosonicated and pasteurized samples showed no significant (p > 0.05) changes in pH, total soluble solids, and titratable acidity. TS improved the cloud value and color attributes. Furthermore, TS enhanced total phenols (10 to17%), flavonoids (5 to 25%), antioxidant activity (10.45% to 14.55%) and retention of ascorbic acid (61.98–83.32%) relative to control. Thermosonicated sample at 80 °C/10 min gives the maximum inactivation of Pectin methyl esterase (PME), Peroxidase (POD) and Polyphenol oxidase (PPO) enzymes. While both thermosonication and pasteurization drastically decreased the microbial count to undetectable levels, only TS exhibited modest improvement in sensory qualities. The results demonstrated that TS can enhance the overall safety, quality, and commercial viability of guava juice as a practical substitute to pasteurization.

Impact of Thermo-sonication on Pectin Methylesterase (PME) Inactivation & Cloud Stability of Nagpur Mandarin Juice

Nagpur mandarin is well known globally for its excellent nutritional benefits and flavour. These all characteristics get affected during storage due to a lack of knowledge & processing techniques. One of the most important parameters which decide the quality of fruit juice during storage is cloud stability, which gets affected during storage due to the action of an enzyme named pectin methylesterase. Pectin methylesterase enzyme (PME) is a heat-resistant enzyme that impacts the juice quality by decreasing the cloud stability. Thermosonication is a technique to reduce the PME activity in juice by utilizing the principle of heat and ultrasound. Therefore, an experiment was carried out to inactivate the PME utilizing the ultrasound combined with higher temperature known as thermosensation on Nagpur mandarin (Citrus reticulata Blanco) juice. This study was initiated to calculate the effect of thermosensation treatment on the pectin methylesterase enzyme (PME) activity along with the effect of thermosensation on cloud stability in comparison with various thermal pasteurization treatments. Juice samples were thermosonicated at temperatures of 50 ℃, 55 ℃, 60 ℃, and 63 ℃ for 25, 20, 15 and 10 min respectively at a constant frequency of 40 kHz. Juice samples were also thermally pasteurized at 65 ℃, 75 ℃, 85 ℃, and 95 ℃ for 80, 60, 40, and 20 sec. Samples were analyzed after treatment at 0 days and 7 days of refrigerated storage at 4 ℃. Result indicated that thermosensation at 63 ℃ for 10 min at 40 kHz frequency showed much reduced PME activity with inactivation of 72.56% over control whereas thermal pasteurization at 95 ℃ showed inactivation of 69.31%. The same treatment also showed maximum cloud activity as compared to control and thermally pasteurized juice samples. Hence thermosensation can be used as an alternative processing technique for the inactivation of PME and cloud stability.

The combined effect of supercritical carbon dioxide and mild temperatures on pectin methylesterase inactivation in orange juice.

Pectin methylesterase (PME) is the target-enzyme in orange juice processing, and its inactivation preserves the original juice's cloud, a valuable quality attribute for consumers. This study was primarily undertaken to evaluate the combination of supercritical carbon dioxide (SC-CO2) and mild temperatures on PME inactivation in the juice. Physicochemical and color parameters were also evaluated. Pressures (P) in the range of 74 to 351 bar, temperatures (T) between 33°C and 67°C, and holding times (t) varying from 20 to 70 min were tested using a central composite rotational design. Determination of PME activity, pH, titratable acidity, soluble solids, ratio, lightness, °hue and chroma in freshly extracted/raw and treated juice samples, and total color difference (TCD) between raw and treated juice were carried out. The percentage of PME reduction widely ranged from 13.7% to 91.8%. The variable P had a significant effect (p ≤ 0.05) on TCD and acidity variation. T exhibited a significant effect on the PME reduction, TCD, and soluble solids variation; however, t impacted on the soluble solids variation only. The interaction between P and t (P.t) affected the soluble solids and acidity variation, and T.t affected TCD. The combination of SC-CO2 and mild temperatures is potentially capable of inactivating PME in orange juice.

Pulsed light treatment of table grape juice: Influence of matrix pH on microbial and enzyme inactivation kinetics

The influence of pulsed light (PL) treatment and matrix pH was tested in the case of green table grape juice. Escherichia coli, Listeria monocytogenes, and Saccharomyces cerevisiae were inoculated into the table grape juices with matrix pH levels of 3.0, 3.5, and 4.0. Further, the juice was treated at 378–3186 J/cm2. The Weibull model better explained the microbial inactivation data. The shape parameters (β) from the Weibull model for E. coli, L. monocytogenes, and S. cerevisiae are 0.95 ± 0.02, 0.55 ± 0.04, and 0.40 ± 0.03, respectively. A lesser δ-value (scale parameter in J/cm2) was obtained for each microorganism at a lower pH. PL exposure resulted in a lack of cell visibility, cell wall, and cell membrane disruption in bacterial cells. The inactivation of polyphenol oxidase (PPO), peroxidase (POD), and pectin methyl esterase (PME) was studied at an elevated fluence level of 1152–3186 J/cm2. Enzymes in the juice were less sensitive to PL than microorganisms. The enzyme inactivation of >90% was achieved at 3186 J/cm2; thus, the juice was adjudged as the microbially safe and enzymatically stable pasteurized sample. The PL-induced enzyme inactivation followed nth-order kinetics with an inactivation order (n) of 1.1, 1.15, and 0.85 for PPO, POD, and PME, respectively. The enzyme inactivation rate was higher at a low pH. The PL-treated pasteurized juice at pH 3.5 showed the highest sensory acceptance (7.4 out of 9), with a minimal color change, a 7% loss in total phenolics, and a 12% loss in vitamin C and flavonoids.

Inactivation of pectinmethylesterase in fresh orange juice by cold atmospheric plasma technology: A kinetic study

Preservation of fruit juices requires the inactivation of natural endogenous enzymes, such as pectinmethylesterase (PME). Within this work, cold atmospheric plasma (CAP), and in particular a dielectric barrier plasma jet fed with helium gas, was demonstrated to effectively inactivate PME of freshly squeezed orange juice in short treatment times (2–30 min). By a combination of temperature measurements and a multidimensional heat transfer model, the temperature profile of the whole sample during plasma treatment was extracted. It was found that the thermal phenomena were not a driving factor for PME inactivation. Plasma treatment of orange juices resulted in inactivation of 55–80% of PME with <5.0% of PME inactivation caused by the temperature increase from 20 to 90 °C. The Weibull distribution model compared to the first-order fractional, the sigmoidal logistic and the Hulsheger's kinetic models was found to better describe mathematically (R2 > 0.99; Af = 1.002–1.052) the effect of CAP processing on residual PME activity. Multi-parameter equation fits allowed the prediction of residual PME activity as a function of the applied voltage, helium flow, and treatment time. Generally, higher voltages and lower helium flows applied led to higher PME inactivation rates in fresh orange juice.

Ultrasound processing of sweet lime juice: Effect of matrix pH on microbial inactivation, enzyme stability, and bioactive retention

AbstractThe effect of pH on microbial, enzyme inactivation, and bioactive retention in ultrasound (US) treated sweet lime (Citrus limetta) juice was studied. The pH of the juice varied at 3.5, 4.0, and 4.5, whereas the US duty cycle (DC) ranged between 75% and 95% (0.07–0.7 W/cm3), and the treatment time varied between 15 and 30 min. More than 5‐log reduction in Saccharomyces cerevisiae and Escherichia coli was achieved at DC 95% (25 min) while Listeria monocytogenes got inactivated at DC 85% (25 min). 95%–99% inactivation of peroxidase and pectin methylesterase was achieved at DC 95% (30 min), while 60% inactivation in polyphenol oxidase (PPO) was observed. The addition of ascorbic acid (1 g/L) as an EI resulted in the complete inactivation of PPO at DC 85% (25 min). A maximum color change (ΔE*) of 2.6 was noticed at the highest US treatment condition (DC 95%, 30 min). Eventually, pH showed a marginal effect (˂5%) on retaining vitamin C and bioactive compounds, but US treatment enhanced them. US treatment at DC 95% (30 min) produced microbially safe (˃5‐log cycle reduction) and enzymatically stable (99% enzyme inactivation) juice. To conclude, optimized US treatment increased total phenolic content (TPC), antioxidant capacity, and vitamin C in sweet lime juice by 16.7%, 10.7%, and 14.1%, respectively.Practical applicationsConsumer demand for superior quality sweet lime juice with minimal processing. This explores the opportunity of applying US technology as one of the nonthermal treatments for such a product. Furthermore, the treated juice should be microbially safe and enzymatically stable with better bioactive preservation. Typically, the pH of the fruit juice is one of the main influencing factors controlling microbiological spoilage. The current study explores the consequence of matrix pH and US conditions on the microbial inactivation, enzyme stability, and bioactive compounds of US‐treated sweet lime juice. The hypothesis is that the ultrasonic power required to achieve microbial safety at a lower pH of sweet lime juice should be lower. This will eventually help the industry choose an optimal harvesting condition to obtain the desired pH during processing.

Effect of High Hydrostatic Pressure and Pulsed Electric Fields Processes on Microbial Safety and Quality of Black/Red Raspberry Juice.

Black and red raspberries are fruits with a high phenolic and vitamin C content but are highly susceptible to deterioration. The effect of high hydrostatic pressure (HHP 400–600 MPa/CUT-10 min) and pulsed electric fields (PEF, frequency 100–500 Hz, pulse number 100, electric field strength from 11.3 to 23.3 kV/cm, and specific energy from 19.7 to 168.4 kJ/L) processes on black/red raspberry juice was studied. The effect on the inactivation of microorganisms and pectin methylesterase (PME) activity, physicochemical parameters (pH, acidity, total soluble solids (°Brix), and water activity (aw)), vitamin C and phenolic compounds content were also determined. Results reveal that all HHP-treatments produced the highest (p < 0.05) log-reduction of molds (log 1.85 to 3.72), and yeast (log 3.19), in comparison with PEF-treatments. Increments in pH, acidity, and TSS values attributed to compounds’ decompartmentalization were found. PME activity was partially inactivated by HHP-treatment at 600 MPa/10 min (22% of inactivation) and PEF-treatment at 200 Hz/168.4 kJ/L (19% of inactivation). Increment in vitamin C and TPC was also observed. The highest increment in TPC (79% of increment) and vitamin C (77% of increment) was observed with PEF at 200 Hz/168.4 kJ/L. The putative effect of HHP and PEF on microbial safety, enzyme inactivation, and phytochemical retention is also discussed in detail. In conclusion, HHP and PEF improve phytochemical compounds’ content, microbial safety, and quality of black/red raspberry juice.

Inactivation of Endogenous Pectin Methylesterases by Radio Frequency Heating during the Fermentation of Fruit Wines

Pectin methylesterase (PME) is a methyl ester group hydrolytic enzyme of either plant or microbial origin. Importantly, endogenous PMEs in fruits can catalyze the demethoxylation of pectin with a bulk release of methanol, largely impacting the fruit juice and wine industries. Here, we demonstrated radio frequency (RF) heating for inactivation of endogenous PMEs and investigated the relevant mechanisms underpinning enzymatic inactivation. The RF heating curve indicated that the optimal heating rate was achieved at an electrode gap of 90 mm (compared to 100 mm and 110 mm) and that the inactivation rate of the enzyme increases with heating time. RF heating exhibited better effects on enzymatic inactivation than traditional water heating, mainly by changing the secondary structures of PMEs, including α-helix, β-sheet, β-turn, and random coil. Moreover, fluorescence spectroscopy indicated changes in the tertiary structure with a significant increase in fluorescence intensity. Significantly, application of RF heating for inactivation of PMEs resulted in a 1.5-fold decrease in methanol during the fermentation of jujube wine. Collectively, our findings demonstrated an effective approach for inactivating endogenous PMEs during the bioprocesses of fruits.

Sonoprocessing of freshly squeezed orange juice: Ascorbic acid content, pectin methylesterase activity, rheological properties and cloud stability

Ultrasound performance at low energy densities (9–86 J/mL) on the quality attributes of freshly squeezed orange juice was compared to heat treatment (90 °C/1 min). Mild sonoprocessing improved orange juice's kinetic stability preserving its ascorbic acid (vitamin C) content. The ultrasound energy densities could not promote the inactivation of pectin methylesterase (PME) in the orange juice samples. However, higher energy densities contributed to their cloud stability along cold storage. In contrast, the heat processing inactivated 96.3% of PME, but thermally treated samples lost their cloudiness and had their ascorbic acid content reduced by 7%. Cloud stabilization after sonoprocessing was mainly attributed to acoustic cavitation performance to modulate particle size distribution. Ultrasound processing is a promising technique to solve one of the main technological issues of the freshly squeezed orange juice. However, mild sonoprocessing must be combined with other innovative non-thermal technologies to provide a safe and high-quality orange juice.

Effect of different microwave power levels on inactivation of PPO and PME and also on quality changes of peach puree

The effect of microwave (MW) treatment with different power densities (4.4, 7.7, and 11.0 W/g) on polyphenol oxidase (PPO) and pectin methyl esterase (PME) inactivation in peach puree were studied, and the changes in color, rheological properties, total polyphenol and flavonoid and antioxidant capacity were evaluated. By using time/temperature data collected during MW heating, three cook values levels (0.36, 10, 24 min) for each power density were calculated. The PPO was significantly decreased from ca. 50% to ca. 5% when increasing the cook value level, regardless of power density applied. While PME significantly decreased from 40.6% to 10.2% when power density increased from 4.4 to 11.0 W/g at cook value 24 min. MW treatment did not alter the flow behaviour of peach puree. The apparent viscosity values of peach puree significantly increased after MW treatment with increasing cook value, regardless of power density applied. The L* values of peach puree significantly increased from 36.98 to 38.10 or more after MW treatment at cook value 10 min and 24 min. MW treatment could maintain the amount of total polyphenol, total flavonoid and antioxidant capacity, preserving the nutritional and functional values of the product.

Effect of thermally assisted hydrodynamic cavitation (HC) processing on physical, nutritional, microbial quality, and pectin methyl esterase (PME) inactivation kinetics in orange juice at different time and temperatures

Orange juice is the second most commonly consumed juice worldwide. However, its quality is deteriorated because pectin methyl esterase (PME), a thermostable enzyme. In this study, orange juice was processed by hydrodynamic cavitation (HC) at 4 bar pressure with varying temperatures (40℃–70℃) and process time (0–8 min). pH and TSS of the juice remained stable during HC processing. A decreased flow behavior index and increased flow consistency index was observed along with sedimentation index, indicating the effectiveness of HC in modifying the physical stability of juice. More retention of vitamin-C, phenolics, and antioxidants was observed in HC-processed juice compared with unprocessed; however, degradation was recorded at higher temperature. Maximum PME inactivation (65%) was observed at 70℃ for 8 min. Microbial inactivation at lower temperatures and time was achieved. Based on this research, it can be concluded that HC could be used as a potential liquid food-processing technique for improving the physicochemical and nutritional quality of orange juice. Practical applications Hydrodynamic cavitation (HC) is an upcoming novel nonthermal processing used for improving the physicochemical, nutritional, and microbial quality of fruit juices. HC is advantageous over other methods because of its low capital cost, reduced time, high efficiency, and lower temperature that help in retention of bioactives in foods. HC also causes inactivation of enzymes and microorganisms, thus improving quality and safety of final product. This novel processing is not yet being explored much and there is huge scope for its application in preservation and processing of different fruit juices and other liquid foods (milk, cereal drinks etc.)

The optimal time-temperature conditions for orange juice microwave−assisted pasteurization

In this work, the inactivation kinetics of pectin methylesterase (PME) and degradation kinetics of ascorbic acid (AA) of batch microwave-assisted pasteurized orange juice were studied, at time-temperature conditions from (70–90) °C between (0 and 180) s. The two-component first-order and the fractional kinetic models were adjusted for PME and AA data, respectively. The microwave treatment was effective for the inactivation of PME with minimal AA retention of 85%. The adjusted kinetics parameters at Tref = 90 °C, for PME were: D1, 90 °C = 0.18 s, D2, 90 °C = 32. 8 s, z1 = 10.6 °C, z2 = 11.8 °C, with α = 0.91. For AA were: k90 °C = 0.02 s−1 and z = 55.4 °C with AA∞/AA0 = 0.85. A time-temperature diagram created from the kinetics parameters indicates the best isothermal conditions for orange juice from 78 s at 82.5 °C to 18 s at 90 °C.

Partial purification, characterization and kinetics of thermal inactivation of pectin methylesterase and polygalacturonase enzymes from Indian lemon (Citrus limon (L.)

The extraction, purification, and characterization of PME and PG were studied for lemon fruit. Enzymes were partially purified by anion-exchange chromatography using UNOsphere™ Q anion exchanger for lemon PME and DEAE Cellulose anion exchanger for lemon PG. A single band with a fold purity of 20.95 and 15.75 for PME and PG respectively was observed on SDS-PAGE. Molecular weights of partially purified fractions of PME and PG as exhibited by SDS-PAGE were 48 kDa and 95 kDa respectively. Optimum pH and temperature of partially purified lemon PME and PG were pH 9, pH 5 and 45 °C. The crude PME and PG extracts exhibited D-values of 120.57–63.79 and 185.72–29.18 min; Z-values of 15.11 and 16.50 °C; and activation energies (Ea) of 58.23 and 57.18 kJ/mol, respectively. The enzyme stability was not significantly influenced by the fruit matrix as reflected by Z and Ea values. D and Z-values of both the enzymes and microorganisms should be considered while designing thermal treatments to obtain a product with better shelf life.

High pressure and thermal processing on the quality of zucchini slices

In response to the market demand for low processed vegetables, high-pressure treatments (400,600 MPa; 1,5 min) were applied on zucchini slices and compared to a traditional blanching treatment. Histological observations, texture and color analysis, pectinmethylesterase (PME) and antioxidant (DPPH) activities were measured and compared to untreated samples. The histological observations revealed that the longer high-pressure treatments (5 min) led to more extended cell lysis and dehydration than the shorter ones (1 min) and blanching. High-pressure treatments resulted less effective than blanching on PME inactivation, with the best results obtained at 400 MPa for 1 min. Comparable texture parameters were observed for high-pressured and blanched samples. The negative correlation found between PME activity and the texture parameter ‘distance of the first peak force’ revealed an effect of PME on the texture recovery after treatments. High pressure led to a general browning of zucchini parenchyma and to DPPH drop. The correlations found between DPPH and color suggest the common nature of the phenomena. The influence of pressure and time on the studied parameters was revealed by two-way ANOVA. Principal component analysis clustered together the four high-pressure-treated samples, being clearly divided by blanched and untreated ones.