Effects Of Supercritical Carbon Dioxide On The Conformational Transitions Of Pure Enzymes And Activity Inactivation Of Three Enzymes In Cantaloupe Juice

As a novel non-thermal process, SCCO2 technology has acctracted more attention. The effects of SCCO2 on the activity and structural denaturalization of Peroxidase(POD), Polyphenol oxidase(PPO) and Lipoxygenase(LOX) were investigated. Gel electrophoresis, far UV-circular dichroism(CD) and tryptophan fluorescence spectropy were used to analysis the primary, secondary and tertiary structure and the configuration of POD, PPO and LOX were observed by TEM. The effects of SCCO2 on the activity of three kinds of enzymes after treatment and during storage period in cantaloupe juice, and at the same time the effects of SCCO2 on color and browning degree were alse investigated. The main conclusions were summarized as follows:(1) Compared with only heat-treatment, SCCO2 had significant effects on sterilization of POD, PPO and LOX, the activity of POD decreased to 44.62% when exposed to the heat-treatment at 65℃for 60 minutes, while, after the SCCO2 treatment at 35MPa with 65℃for 60 minutes, the residual activity of POD was 1.96%. The activity of PPO decreased to 18.09% when exposed to the heat-treatment at 65℃for 60 minutes, while, after the SCCO2 treatment at 15MPa with 55℃for 30 minutes, the residual activity of PPO was 0.09%, when processing time was 45 minutes, the activity was 0. The activity of LOX decreased to 26.78% when exposed to the heat-treatment at 65℃for 60 minutes, while, after the SCCO2 treatment at 15MPa with 55℃for 5 minutes, the residual activity of PPO was 0.22%, when processing for 15 minutes, the activity of PPO was completely inactive.(2) SDS-PAGE indicated that no breakage of peptide and no primary structure of POD, PPO and LOX protein occurred, and it illustrated that the Amino acids'sequence didn't destroyed. TEM also found the diameter of POD, PPO and LOX molecules became larger, which account for the aggregation of POD, PPO and LOX molecules possibly.(3) The CD and tluorescence spectrum were changed after SCCO2 treatment. Theα-helix relative content in the secondary structure decreased and the intrinsic relative fluorescence intensity increased as the pressure of SCCO2 treatment was elevated.(4) SCCO2 treatment has a good inactive effects to POD, PPO and LOX in cantaloupe juice, analysis of first-order reaction kinetic data showed that D values of POD, PPO and LOX in cantaloupe juice were closely to the pressure and temperature of SCCO2 treatment. Higher pressures and higher temperatures resulted in lower D values and higher k values. The activity of POD, PPO and LOX in cantaloupe juice had different degree recovery during storage at 4℃for 4 weeks, and the degree of the recovery had a relation with treatment condition such as pressure. The recovery degree of the activity of POD, PPO and LOX in cantaloupe juice had a relationship with treatment condition such as pressure, the samples with higher pressure and longer treatment time had less recovery degree. (5) After SCCO2 treatment, the L value in cantaloupe juice decreased, and the degree of the decline is the least at 35MPa.ΔE value increased by 1.15 at the condition of 35MPa, the increased degree is the least, and the change of colour in cantaloupe juice is most not obvious. With the pressure elevated, treatment time is enlonged, and it could inhibited the browing of the cantaloupe juice effectively. The A values in treated samples were less than the untreated samples obvious. SCCO2 had a better inhibitory effect to the colour and browing degree of the cantaloupe juice, and the greater of the pressre, the better of the inhibitory effect.