Modelling the effect of superatmospheric oxygen concentrations on in vitro mushroom PPO activity

The kinetics of polyphenol oxidase (PPO, EC 1.14.18.1) with respect to oxygen concentrations from 5 to 100% using chlorogenic acid (CGA) as substrate was examined. In vitro mushroom PPO activity was determined by measuring the consumption of oxygen during the oxidation reaction. A differential Michaelis–Menten model was fitted to the obtained total depletion curves. The product concentration as well as the concentration of oxygen had a clear inhibitory effect on the reaction rate. However, the inhibitory effect of oxygen was more evident at low product concentration. A linear mixed inhibition model that considered both the product (oxidised CGA) and oxygen as inhibitors was developed. A model with the product as a competitive inhibitor and oxygen as an uncompetitive inhibitor was the most appropriate to explain the reaction kinetics. The values of the inhibition constants calculated from the model were 0.0032 mmol L⁻¹ for K_m (Michaelis–Menten constant related to oxygen), 0.023 mmol L⁻¹ for K_mc (constant for competitive inhibition due to the product), 1.630 mmol L⁻¹ for K_mu (constant for uncompetitive inhibition due to oxygen) and 1.77 × 10⁻⁴ mmol L⁻¹ s⁻¹ for V_max (maximum reaction rate). The results indicate that superatmospheric oxygen concentrations could be effective in preventing enzymatic browning by PPO. Copyright © 2006 Society of Chemical Industry     

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

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

Purification and enzymatic characteristics of a novel polyphenol oxidase from lotus seed (Nelumbo nucifera Gaertn.)

A polyphenol oxidase (PPO) from lotus seed was purified by the procedures including ammonium sulphate precipitation and affinity chromatography. The apparent molecular mass was 38.6 kDa by SDS‐PAGE. Kinetic studies showed that the K_m and V_max values for catechol were 6.04 mm and 416.67 U, respectively. The PPO performed optimal activity in 20 °C and pH 7.0. The enzymatic activity could be mainly maintained up to 50 °C and pH 4.0–7.0. The activity could be inhibited by various inhibitors including thiourea, urea, sodium hydrogen sulphite, EDTA·2Na, SDS, citric acid, guanidine hydrochloride, ascorbic acid, sodium sulphite and sodium thiosulphate. The metal ions Ba²⁺, Mg²⁺, Ca²⁺, Mn²⁺, Co²⁺ and Zn²⁺ could inhibit the activity of PPO, while Cu²⁺ performed obvious enhancement. The enzymatic properties of PPO could probably provide practical application in inhibiting the PPO activity and preventing enzymatic browning in the process of picking, transportation, processing and storage of fresh lotus seeds.     

双孢蘑菇酶促褐变特性及褐变的控制

为有效控制双孢蘑菇褐变,研究pH值、温度、底物浓度以及抑制剂对双孢蘑菇的多酚氧化酶(polyphenol oxidase,PPO)活性的影响。结果表明：以邻苯二酚为底物,双孢蘑菇PPO最适反应pH6.8,最适反应温度20℃,最适反应底物浓度为0.06mol/L,米氏常数Km为0.1072mol/L。采用计算机图像处理技术对双孢蘑菇褐变程度进行定量检测,得出对双孢蘑菇贮藏保鲜效果较好的褐变抑制剂及其最佳配比为0.30mmol/L半胱氨酸、0.06％抗坏血酸、0.05%乙酸。     

Purification of polyphenol oxidase and the browning control of litchi fruit by glutathione and citric acid

Polyphenol oxidase (PPO, EC 1.10.3.2) was purified to homogeneity from litchi peel yielding a single protein with a molecular weight of about 75.6 kD by Sephadex G‐100 gel filtration, and a 108‐fold purification of PPO achieved. The enzyme was determined to be composed of two similar subunits. Glutathione, L ‐cysteine and citric acid suppressed PPO activity markedly, whereas ascorbic acid and n‐propyl gallate showed a little inhibition. Moreover, the effect was enhanced by the addition of citric acid. On the basis of the inhibition of PPO activity in vitro, the use of 10 mmol l ⁻¹ glutathione and 100 mmol⁻¹ l citric acid was found to give good control of the browning of litchi fruit, and an 80–85% inhibition of PPO activity was observed. It is suggested that application of glutathione in combination with citric acid may slow down the browning of litchi fruit. © 1999 Society of Chemical Industry     

Prevention of Enzymatic Browning of Chinese Yam (Dioscorea spp.) Using Electrolyzed Oxidizing Water

In this study, the effects of electrolyzed oxidizing water (EOW) on the prevention of enzymatic browning of fresh‐cut “Jiu Jinhuang” Chinese yam were investigated. The yams were immersed in the inhibitors for 25 min at 20 °C. Compared with the tap water (TW) treatment, the chromatic attributes were significantly different after 72 h of storage (P < 0.05). The activities of polyphenol oxidase (PPO, EC 1.10.3.1), peroxidase (POD, EC 1.11.1.7), and _L‐phenylalanine ammonia lyase (PAL, EC 4.3.1.5) were inhibited when measured at 24 h. The contents of phenolic acids, including gallic and chlorogenic acid, in the group treated with the slightly acidic electrolyzed water (SAEW) were higher than those treated with TW and neutral electrolyzed water (NEW). The group treated with NEW had the highest total phenol content (P < 0.05, at 24 h), while the group treated with SAEW had the highest flavonoid content (P < 0.05) during storage. Without being treated with inhibitors, the K_m and V_max values of yam PPO were 0.0044 mol/L and 0.02627 U/min, respectively, and the K_i of samples treated with SAEW and citric acid (CA) were 15.6607 and 2.3969 μmol/L, respectively. These results indicate that EOW is beneficial as a browning inhibitor.     

Partial characterization of lettuce (Lactuca sativa L.) polyphenol oxidase

Polyphenol oxidase (PPO) from garden lettuce (Lactuca sativa L.) was partially purified by ammonium sulphate ((NH₄)₂SO₄) precipitation and dialysis, and then some of its kinetic properties such as optimum pH and temperature, substrate specificity, thermal inactivation and inhibition were investigated. The total phenolic and protein contents of Lactuca sativa L. extracts were determined according to the Folin-Ciocalteu and Bradford methods, and found to be 304 mg/100 g on a fresh weight basis and 494 μg/mL, respectively. PPO activity was determined using 4-methylcatechol, catechol and pyrogallol as substrates. Kinetic parameters, K _m and V _max, were calculated from Lineweaver–Burk plots. According to V _max/K _m ratio, pyrogallol was the most suitable substrate, followed by catechol and 4-methylcatechol. The optimum temperature and pH values were 30, 40 and 30 °C; and 6.5, 8.0 and 7.5 for 4-methylcatechol, catechol and pyrogallol substrates, respectively. The thermal inactivation of PPO was investigated at 35, 55 and 75 °C. The enzyme activity decreased with increasing temperature. The effect of different inhibitors on partially purified Lactuca sativa L. PPO was spectrophotometrically investigated. For this purpose, tropolone, glutathione, ascorbic acid and 4-aminobenzoic acid were used to inhibit the activity of Lactuca sativa L. PPO at different concentrations. From the experimental results, it was found that glutathione was found to be the most potent inhibitor for Lactuca sativa L. PPO.     

Determination of some biochemical properties of polyphenol oxidase from Emir grape (Vitis vinifera L. cv. Emir)

Polyphenol oxidase (PPO) was extracted from Emir grapes grown in Turkey and its characteristics in terms of pH and temperature optima, thermal inactivation, kinetic parameters and potency of some PPO inhibitors were studied. The optimum pH and temperature for grape PPO were found to be 4.2 and 25 °C respectively using catechol as substrate. K_m and V_max values were found to be 25.1 ± 2.72 mmol L⁻¹ and 0.925 ± 0.04 OD₄₁₀ min⁻¹ respectively. Of the inhibitors tested, the most potent was sodium metabisulfite, followed by ascorbic acid. The thermal inactivation curve was biphasic. Activation energy (E_a) and Z values were calculated as 251.4 kJ mol⁻¹ (r² = 0.996) and 8.92 °C (r² = 0.993) respectively. Copyright © 2006 Society of Chemical Industry     

BROWNING INHIBITION AND QUALITY MAINTENANCE OF FRESH-CUT CHINESE WATER CHESTNUT BY ANOXIA TREATMENT

Experiments were conducted to test the effects of anoxia treatments on the quality of fresh‐cut Chinese water chestnut (CWC). Fresh‐cut CWC was exposed to pure N₂ for 0 (control) or 4 h, then placed in trays overwrapped with plastic film and finally stored at 4C. Changes in surface discoloration, eating quality, disease incidence, concentrations of total soluble solids and titratable acidity, activities of phenylalanine ammonia lyase (PAL) and polyphenol oxidase (PPO), and membrane permeability were analyzed. The short‐term anoxia treatment significantly inhibited surface discoloration and disease development, and markedly reduced the loss of eating quality in association with a higher concentration of total soluble solids. Exposure of fresh‐cut CWC to pure N₂ gas for 4 h significantly reduced membrane permeability, but it increased PAL and PPO activities. Thus, the browning inhibition of the fresh‐cut CWC by the short‐term anoxia treatment may be because of the maintenance of compartmentation between enzymes and their substrates, preventing enzymatic reaction. The result suggests that short‐term anoxia treatment exhibits potential for extending the shelf life of fresh‐cut CWC.     

Characterization of polyphenol oxidase from Uapaca kirkiana fruit

Polyphenol oxidase (PPO), the enzyme responsible for the postharvest spoilage of fruits, was extracted and purified from Uapaca kirkiana peel and pulp by ammonium sulfate precipitation and dialysis. Further purification of peel PPO was carried out by gel filtration chromatography. Optimum pH values were 7 and 8 for peel and pulp PPO, respectively. The optimum temperatures for peel and pulp PPO were 45 and 35 °C, respectively. Inhibition studies of the PPO enzyme were performed using citric acid, sodium azide, sodium metabisulfite and thiourea. The most effective inhibitors were sodium azide and citric acid for both peel and pulp PPO. V_max and K_m values were 13.63 units min^?1 and 4.923 mmol L^?1, respectively, for peel PPO and 14.03 units min^?1 and 5.43 mmol L^?1, respectively, for pulp PPO. Three isoenzymes of Uapaca kirkiana PPO were detected by polyacrylamide gel electrophoresis. One of the isoenzymes could be identified as having a molecular weight of 26 625 Da. Copyright © 2005 Society of Chemical Industry     

Some properties of polyphenol oxidase from lily

A study of crude polyphenol oxidase (PPO) from lily bulbs was carried out to provide information useful for guiding food processing operations. Optimum pH for the enzyme activity in the presence of catechol, were 4.0 and 7.0 at room temperature(approximately 20 °C) and the enzyme was stable in the pH range from 5.0 to 6.5 at 4 °C for 10 h. Its optimum temperature was 40 °C and the heat inactivation of the enzyme followed first‐order kinetics. Lily PPO possessed a diphenolase activity toward catechol, catechin and gallic acid; catechin was the best substrate for the enzyme considering the V_max/K_m ratio. The most effective enzyme inhibitor was sodium sulphite, although ascorbic acid, l ‐cysteine and thiourea were also effective inhibitors at high concentration. But NaCl and citric acid were poor inhibitors of the enzyme. Data generated by this study might help to better prevent lily bulbs browning.     

Polyphenoloxidase from DeChaunac grapes

Polyphenoloxidase (PPO) from red grape cultivar, DeChaunac, grown in New York State was isolated and purified 17-fold by using Phenyl Sepharose CL-4B column. Disc gel electrophoresis revealed near homogeneity of three isoenzyme bands. The molecular weight of this enzyme ranged from 73 000 to 85 000. The temperature and pH optima of the purified enzyme were 20 °C and 6.0, respectively. Kinetic studies showed that the thermal inactivation of the PPO followed first-order kinetics, with the activation energy, Ea = 52.39 Kcal mol^?1. The substrate specificity showed a high degree of PPO activity toward o-diphenolic compounds with the highest affinity toward caffeic acid among substrates studied. The apparent K_m values for caffeic acid and 4-methylcatechol as substrates for the Dechaunac PPO were 16 mmol and 25 mmol, respectively. The most potent inhibitors of the PPO were D.L-dithiothreitol and sodium metasulphite at the concentration level of 0.5 mmol.     

Polyphenol oxidase activity,phenolic acid composition and browning in cashew apple (Anacardium occidentale, L.) after processing

This study describes the extraction and characterisation of cashew apple polyphenol oxidase (PPO) and the effect of wounding on cashew apple phenolic acid composition, PPO activity and fruit browning. Purification factor was 59 at 95% (NH₄)₂SO₄ saturation. For PPO activity, the optimal substrate was catechol and the optimum pH was 6.5. PPO K_m and V_max values were 18.8 mM and 13.6 U min⁻¹ ml⁻¹, respectively. Ascorbic acid, citric acid, sodium sulphite and sodium metabisulphite decreased PPO activity, while sodium chloride increased PPO activity. Wounding at 2 °C and 27 °C for 24 h increased PPO activity but storage at 40 °C reduced PPO activity. Gallic acid, protocatechuic acid and cinnamic acid (free and conjugate) were identified in cashew apple juice. Cutting and subsequent storage at 40 °C hydrolysed cinnamic acid. 5-Hydroxymethylfurfural content in cashew apple juice increased after injury and storage at higher temperatures, indicating non-enzymatic browning.     

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

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

Characterization and inhibition of <Emphasis Type="Italic">Rosmarinus officinalis</Emphasis> L. polyphenoloxidase

Polyphenoloxidase (PPO) from Rosmarinus officinalis L. was fractionated by ammonium sulfate ((NH₄)₂SO₄) precipitation and dialysis, and then some of its kinetic properties such as optimum pH and temperature, substrate specificity, thermal inactivation, and inhibition were investigated using 4-methylcatechol, catechol, and pyrogallol as substrates. The protein content of Rosmarinus officinalis L. extracts was determined according to Bradford’s method. Kinetic parameters, K _m and V _max, were calculated from Lineweaver–Burk plots. According to V _max/K _m ratio, 4-methylcatechol was the most suitable substrate. The optimum temperature and pH values were 20, 30 and 30 °C, and 7, 8 and 8 for 4-methylcatechol, catechol, and pyrogallol substrates, respectively. The thermal inactivation of PPO was investigated at 35, 55, and 75 °C. The enzyme activity decreased with increasing temperature. The effect of different inhibitors on partly purified Rosmarinus officinalis L. PPO was spectrophotometrically investigated. For this purpose, ascorbic acid and l-cysteine were used to inhibit the activity of Rosmarinus officinalis L. PPO at different concentrations. From the experimental results, it was found that l-cysteine is a more effective inhibitor than ascorbic acid due to lower K _i values.     

Characterization of Sultaniye grape (Vitis vinifera L. cv. Sultana) polyphenol oxidase

Polyphenol oxidase (PPO) was extracted from Sultaniye grapes grown in Turkey, and its characteristics in terms of pH and temperature optima, thermal inactivation, kinetic parameters and potency of some PPO inhibitors were studied. Optimum pH and temperature for grape PPO were found to be 3.4 and 30 °C, using catechol as substrate. K_m and V_max values were found to be 44.5 ± 5.47 mm and 0.695 ± 0.0353 OD₄₁₀ min^?1, respectively. Four inhibitors were tested in this study and the most potent inhibitor was sodium metabisulphite, followed by ascorbic acid. From the thermal inactivation studies in the range of 65–80 °C, the half‐life values of the enzyme ranged between 2.6 and 49.5 min. Activation energy (E_a) and Z values were calculated to be 208.5 kJ mol^?1 (r² = 0.9544) and 10.95 °C (r² = 0.9517), respectively.     

ning reactions during storage of low-moisture Australian sultanas: Effects of vine nitrogen nutrition on subsequent arginine-mediated Maillard reactions during storage of dried fruit

Ten‐year old Vitis Vinifera L. cv. Sultana (Thompson Seedless) grape vines were transferred into large cement pots in low nitrate soil and supplemented with four levels of nitrogen application in the form of ammonium nitrate as follows: 0 g ( N₀ ), 8.5 g ( N₁ ), 17.0 g ( N₂ ) and 25.0 g ( N₃ ). Grapes were harvested from each nitrogen treatment, dipped and dried to low moisture (?11.4%) and subsequently stored for 10 months at 10^oC and at 30^oC in either the presence or absence of oxygen. The pre‐storage concentration of free arginine, free‐proline and total protein in the dried sultanas increased with soil nitrogen application. Skin‐polyphenoloxidase (PPO) activity was higher in sultanas with added soil nitrogen ( N₁, N₂, N₃ ), however pre‐storage differences in total phenolics, and the PPO substrate, trans‐caftaric acid, were not significantly different. After 10 months storage at 30^oC significant browning was observed in both the presence and absence of oxygen. Greater browning corresponded to higher soil‐nitrogen application rates. The concentration of skin trans‐caftaric acid did not decrease in N₀, N₁ or N₂ after 10 months at 30^oC, in either the presence or absence of oxygen, although some decreases were measured in the highest nitrogen sultanas ( N₃ ). While PPO oxidation of the phenolic substrate trans‐caftaric acid was not the primary route to browning, a number of Maillard reaction products (MRP) were present in both sultanas and arginine‐glucose model systems. Those products were separated via reverse phase HPLC and partially characterised by UV‐diode array spectroscopy. The lack of oxidation of trans‐caftaric acid observed in the nitrogen storage trial was confirmed in an accelerated browning experiment in low‐moisture sultanas, where despite browning at 50^oC after 12 days, no decreases in this primary substrate for PPO‐mediated oxidation were measured.     

Browning reactions during storage of low-moisture Australian sultanas: Further evidence for arginine-mediated Maillard reactions during storage, and some effects of vine-shading and harvest date

Sultana grapevines (Vitis Vinifera L. cv. Sultana syn. Thompson Seedless) were subjected to four shading regimes: 50% shading (1), 25% shading (2), fully exposed‐top of canopy (3) and beneath canopy (4) and harvested early (21 February) and late (13 March) in the 1996/1997 sultana season. Grapes from each of the eight field‐treatment combinations represented a range of maturities (14.4 to 23.50^oBrix). Grape samples from each of the treatments were dipped and dried to 18% moisture, with half of each of the sultana samples further reduced in moisture by sunfinishing on plastic sheets in direct sun. These field treatments resulted in sixteen unique dried sultana bulk samples with a range of initial chemico‐physical properties; a_w (0.481–0.691), skin‐polyphenoloxidase (PPO) activity (4.40–9.05 μmol O₂/g.minute) free arginine in skin tissues (1.0–5.10 mg/g) and protein (16.40–27.18 mg/g). Sultanas were stored at 10^oC and 30^oC in either the presence or absence of oxygen for 10 months, and changes in CIE L*a*b* tristimulus values, hue‐angle (h_ab*) and chroma (C_ab*) were monitored. Significant changes in sultana colour occurred in samples stored at 30^oC, especially in higher a_w non‐sunfinished sultanas. Although browning was more intense in the presence of oxygen, significant browning also occurred in the absence of oxygen. Lower concentrations of 5‐hydroxy methylfurfural, a key marker of Maillard browning in samples stored at 30^oC in the presence of oxygen, indicated that the non‐enzymatic reactions were sensitive to oxygen. Changes in the concentration of trans‐caftaric acid, the main substrate of grape PPO, were also measured during sultana drying. Storage browning (changes in L*, b*, h_ab*, C_ab*)in dried sultanas could be predicted by regression models using pre‐storage a_w, free‐skin arginine or Kjeldahl protein after 10 months' storage between 10^oC and 30^oC. Non‐enzymatic and Maillard‐type reactions (sensitive to both oxygen and a_w), made an important contribution to sultana storage browning. We provide only weak evidence that either shaded (immature) or green fruit was more susceptible to storage browning.     

Isolation and characterisation of trypsin from sardinelle (Sardinella aurita) viscera

BACKGROUND: In Tunisia, sardinelle (Sardinella aurita) catches totalled about 13 300 t in 2002. During processing, solid wastes including heads and viscera are generated, representing about 30% of the original raw material. Viscera, one of the most important by‐products of the fishing industry, are recognised as a potential source of digestive enzymes, especially proteases with high activity over a wide range of pH and temperature conditions. This paper describes the purification procedure and some biochemical characterisation of trypsin from S. aurita viscera. RESULTS: Trypsin from the viscera of sardinelle (S. aurita) was purified by fractionation with ammonium sulphate, Sephadex G‐75 gel filtration, Sepharose mono Q anion exchange chromatography, ultrafiltration and a second Sephadex G‐75 gel filtration, resulting in a 5.42‐fold increase in specific activity and 6.1% recovery. The molecular weight of the purified enzyme was estimated to be 24 kDa using size exclusion chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified enzyme showed esterase‐specific activity on N‐α‐benzoyl‐L ‐arginine ethyl ester (BAEE) that was four times greater than its amidase‐specific activity on N‐α‐benzoyl‐DL ‐arginine‐p‐nitroanilide (BAPNA). The optimal pH and temperature for enzyme activity were pH 8 and 55 °C respectively using BAEE as a substrate. The trypsin kinetic constants K_m and k_cat on BAPNA were 1.67 mmol L^?1 and 3.87 s^?1 respectively, while the catalytic efficiency k_cat/K_m was 2.31 s^?1 L mmol^?1. CONCLUSION: Trypsin was purified from sardinelle (S. aurita) viscera. Biochemical characterisation of S. aurita trypsin showed that this enzyme can be used as a possible biotechnological tool in the fish‐processing and food industries. Copyright © 2008 Society of Chemical Industry     

雪梨多酚氧化酶酶学特性及其果汁褐变控制技术研究

雪梨在加工过程中极易发生酶褐变,为抑制雪梨汁的酶褐变,试验以雪梨为原料制备多酚氧化酶(Polyphenol oxidase,PPO)粗酶提取液,加入邻苯二酚为底物,采用分光光度法对其PPO的酶学特性及不同护色剂对PPO活性的影响进行了研究;并选择L-半胱氨酸、D-异抗坏血酸钠、抗坏血酸3种添加剂对雪梨汁进行护色,通过响应面优化试验确定雪梨汁最优护色组合。结果表明:雪梨PPO的最适pH为4.5,最适温度为30 ℃;雪梨PPO具有一定的热稳定性,随着温度的提高,抑制PPO活性所需要的时间逐渐减少;雪梨PPO催化底物邻苯二酚的酶促反应动力学与米氏方程高度符合,最大反应速率Vmax=217.39 U/min,米氏常数Km=0.152 mol/L。雪梨汁加工的最优护色剂组合为:6.56 mmol/L的L-半胱氨酸、4.58 mmol/L的D-异抗坏血酸钠和6.18 mmol/L的抗坏血酸,在此条件下对雪梨汁褐变的抑制率可达90.82%。