| Recovery of the fish muscle protein isolate by acid/alkaline solubilization/isoelectricprecipitation method, also known as pH-shift process, became promising in fish industry inrecent years. Compared to the traditional surimi process, the pH-shift method for proteinisolate/surimi production has several advantages, including elimination of the mechanicalseparation process of muscle from bones/skin, lower request for raw material, higher proteinrecovery, and that the protein isolates retain capacity to form gels with good quality. ThepH-shift process provides basic technology support for the updating of traditional aquaculture,however, there are still some problems needed to be considered, for example, how to reducethe consumption of alkaline and acid, which is harmful to the environment.In the present study, high intensity ultrasound (HIU) was applied in the acid/alkalinepH-shift process to recover tilapia (Oreochromis niloticus) protein isolate (TPI) from tilapiafillet muscle. The results in the protein solubility, consistency, sediment ratio, proteinrecovery in TPI, and the protein profile all through the pH-shift process as affected by HIUwere evaluated. In addition, the physic-chemical properties, microstructure, gel strength, andgelation process were investigated. The detailed results are as follows.HIU after homogenization or/and after pH-adjustment to alkaline were effective inincreasing the protein solubility (e.g. when HIU was applied both after homogenization andpH-adjustment to pH10.5, protein solubility was increased from85.3%to97.0%), reducingthe sediment ratio after the first centrifugation and the pI protein solubility. Hence, the proteinrecovery was significantly increased. The protein recovery in conventional pH-shift processat pH11.5was62.4%, which is equivalent to the results (62.6%) at pH10.5aided by twoHIUs, indicating HIU could reduce the alkaline consumption (c.a.40%) and thecorresponding acid comsumption for pH readjustment to pI.SDS-PAGE results showed that HIU caused the degradation of titin and released myosinfrom the bind of titin with thick filament. The bind of nebulin with thin filament could also bedisrupted. Hence more actin and tropomyosin were released. After HIU, the pH value of themuscle homogenate was significantly reduced, indicating more amino residues werenegatively charged, which strengthened the electrostatic repulsion. This might be the reason for the improvement on pH-shift process caused by HIU.Aided by HIU, the solubilized protein tent to dissociate and unfold to greater extent inalkaline condition, which resulted in a less ATPase recovery and increased surfacehydrophobicity in the solubilized protein after the protein refolded at neutral condition.Increased surface hydrophobicity might make it easy for the proteins to interact with eachother, hence denser protein isolate formed at pI point. The results of gel texture test showedthat, aided by HIU or increasing the solubilization pH, the gel strength of TPI was wellimproved. According to the results in the dynamic viscoelastic process of TPI, it might beinferred that the protein conformation and the salt solubility of TPI was significantly changedby HIU.For acid pH-shift process, HIU also significantly reduced the consistency (e.g. whenHIU was applied after homogenization and pH-adjustemetn to pH4.0, consistency wasdecreased from430to310mPa·s) of the muscle homogenate, increased the protein solubility(from83.3%to92.4%), decreased the sediment ratio and the protein solubility at pI. As aresult, the protein recovery was significantly elevated (from54.6%to63.0%), implied thatHIU could reduce the consumption of acid for the pH adjustment and the alkalineconsumption in the subsequent readjustment to pI. The results also showed that HIU was notas effective in the acid process as in the alkaline process in improving the pH-shifttechnology.The electrophoresis analysis for the protein profile through the acid pH-shift processrevealed that HIU also prompted the degradation of titin and disruptured the bind of nebulinwith thin filament. This caused more myosin, actin, and tropomyosin et al. to be releasedfrom the sediment and become solubilized. However, compared to the alkaline pH-shiftprocess, HIU exhibited less of an effect in promoting the solubilization of myosin, actin, andtropomyosin et al. in acid pH-shift process.When the same HIU combination treatment was applied, reducing pH (more acidcondition) did not demonstrate effect on the moisture, whiteness and the microstructure ofTPI, which is different from increasing pH value at alkaline condition. Strengthened gelstrength of TPI was obtained in more acid condition. The dynamic viscoelasticity resultsshowed the TPI gelation property was altered as HIU was combined with acid-pH shift process, implying that the protein conformation and the salt solubility of TPI weresignificantly affected by HIU.In the SDS-PAGE results of the TPI gel, no cross-link of myosin was observed, neitherwas protein degradation. This indicated that the improvement in the TPI gel caused by HIU,was mostly due to hydrophobic interaction, which was the results of more exposedhydrophobic clusters in the HIU treated TPI. No cross-link of myosin was observed in the gelfrom TPI, with or without HIU treatment, when setting process was involved in the gelpreparation, implying that most of the endogenous TGase might be separated from the TPIduring the second centrifugation of the pH-shift process, or denaturation possibly occurred tothe endogenous TGase during the acid or alkaline treatment.ATPase activity test, intrinsic tryptophan inflorescence analysis, far-CD, and ANSsurface hydrophobicity test were performed to evaluate the change in conformation of tilapiamyosin as affected by HIU. The results indicated that HIU could cause the slight dissociationof light chain and unfolding of the myosin head at neutral condition. At acid condition, HIUdid not showed effect on the-helix of the myosin, but caused the slight unfolding of themyosin head. At alkaline condition, HIU demonstrated stronger effect on secondary andtertiary structure of myosin than at acid condition: more exposed hydrophobic clusters andmore unfolding of helix. |
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