The Development Of Water-free Transportation For Live Litopenaeus Vannamei And The Biochemical And Muscular Stress Responses

Live pacific white shrimp(Litopenaeus vannamei,shorted as shrimp)are tasty,nutrition-rich and popular among consumers.The shrimp is mainly farmed in the southeast coastal area of China and supplied to the whole country.At present,the main way to achieve the transfer of live shrimp is water transportation,by this way water acts as a culture media of shrimp,the big load of water would inevitably increase transportation costs,and the deterioration of water quality would cause a large number of deaths.This paper proposed a low-cost and efficient strategy of water-free transportation(WFT)for live shrimp.The mechanisms underlying stress resistance and the meat quality changes of shrimps during WFT were studied.The obtained results are listed as followings:(1)After cold acclimation,shrimp are immobilized by a brief cold shock(8°C,3min)in seawater,then packaged in oxygen-filled polyvinyl chloride(PVC)bags and stored at 13°C.The survival rate of shrimp after storage could reach by 96%.Shrimp treated with the“temperature-controlled temporary resting and cold shock device”,the"capture-transport cage"and the"box for water-free live transport"under the above mentioned condition,more than 90%shrimp can survive after long-distance transports(distance>300 km,time>5 h).(2)With the improvement of air exposure tolerance,HSP70 in shrimp hemocytes was upregulated in m RNA and protein levels after cold shock treatment.Both HSP70 RNA interference(RNAi)gene knockdown and recombinant HSP70(r HSP70)injection were successfully established in order to investigate the role of HSP70 in response to air exposure stress.Shrimp receiving r HSP70 showed an improved survival rate(80%)with no significant difference compared to cold shock treated shrimp(control,90%)under air exposure(p>0.05),but the survival rate of HSP70-knockdown shrimp was significantly lower(62%,p<0.05).Reactive oxygen species(ROS)content,relative expression of cytochrome c,caspase-3 activity,and apoptosis rate in hemocytes of HSP70 enriched shrimp(i.e.,cold shock and r HSP70 injection)were significantly lower than HSP70-knockdown shrimp(p<0.05).Results suggested that HSP70 could be induced by cold shock and contributed to improve the tolerance of shrimp suffering air exposure by blocking the apoptosis pathway through scavenging intracellular ROS,inhibiting cytochrome c expression,and inactivating caspase-3.(3)Because of the higher energy demand,air exposure stress during mimicked water-free transportation(WFT)also caused a significant increase in the content of reactive nitrogen species(RNS)in shrimp muscle,and the peroxide value(PV)of the lipids as well as the carbonyl(CO)concentration,intrinsic fluorescence intensity(IFI)and surface hydrophobicity of the proteins in the muscle significantly increased under the oxidation of RNS.However,the SH of the proteins kept away from oxidation as it is only sensitive to reactive oxygen species(ROS).Oxidative damage also leads to degradation of the myosin heavy chain(MHC)and actin.(4)The increments of intracellular Ca²⁺and AMP/ATP ratio activated the AMPK pathway,and thereby promote the consumption of glycogen,accumulation of lactate,and lipolysis under the enzymolysis.Meanwhile,ATP degradation along with glycolysis resulted in the generation of ATP-related adenosine phosphates including IMP.Therefore,the umami taste and freshness of shrimp muscle were improved after WFT.Triglycerides in shrimp muscle are broken down by adipose triglyceride lipase(ATGL),but minor difference in fatty acid compositions and proportions was detected comparing with untreated shrimp.This suggested that glycolysis of muscle glycogen is the main pathway for energy supply of shrimp during WFT.(5)The general proteolytic activity of endogenous proteases significantly increased after WFT.Calpains and collagenolytic proteases were activated due to Ca²⁺overload in the myocytes,and the accumulation of acidic substances such as lactic acid resulted in the increased activity of cathepsins(p<0.05).However,elevated Ca²⁺concentration showed no influence on the activity of the apoptosis-executing enzyme(i.e.,caspase-3)and the apoptosis rate of myocyte.The general proteolytic activity of endogenous proteases was also significantly increased in shrimp from WT due to the increased activity of cathepsin L(p<0.05).The activity of other proteases showed no significant changes compared to the untreated shrimp.(6)During WFT of shrimp,the increased protein surface hydrophobicity caused by oxidation,the degraded intermuscular connective tissue due to the increased collagenolytic proteases activity,and the weakened binding force of proteins to water molecules attributed to the acidified p H,synthetically promoted the conversion of immobilized water into free water in the muscle,which ultimately manifested by the reduced water-holding capacity and increased drip loss of shrimp muscle after WFT.The degradation of myofibrillar proteins in muscle fibers under proteolysis by calpains and cathepsins led to a significant increase in the myofibril fragmentation index(MFI,p<0.05),which indicated the break of muscle fibers during WFT.The break of the muscle fibers and the degradation of the intermuscular connective tissue eventually resulted in the softness of shrimp muscle after WFT(p>0.05).However,because of the SHs of MP were better preserved during WFT,the springiness of shrimp muscle was slightly improved compared to the untreated shrimp(p>0.05).Although the textural properties of the muscles of the shrimps were slightly altered after WFT,there were no significant differences compared to the untreated shrimp(p>0.05).These results suggested WFT would not deteriorate the meat quality of shrimp.Although the degree of oxidation in lipids and proteins was lower,the protease activities were lower,and the metabolic pattern was weaker in shrimp from WT,only the water-holding capacity of the muscles was slightly better than that of shrimp from WFT.