The Effect Of Lysine And Galactose On The Thermal Conversion Of Trimethymine-N-Oxide In Squid And Kinetics Of Model

As a big producer and consumer of aquatic products, China plays a crucial role in aquatic products industry. However, in recent years, an endless stream of seafood safety issues, especially endogenous formaldehyde (FA) problem, severely restricted the export of squid products and even the development of entire aquatic products industry in China. Extensive studies were showed that the precursor of fonnaldehyde in seafoods was trimethylamine N-oxide (TMAO). In aquatic products, FA and dimethylamine (DMA) could be produced from TMAO degradation by TMAOase catalysis and non-enzymatic degradation pathway. At present, the property and activity of TMAOase were studied extensively in various species of marine product. However, the thermal degradation pathway was rarely reported. In view of this, based on the establishment of ion chromatography detection method for DMA, TMA and TMAO, the endogenous components stimulating thermal decomposition of TMAO were investgated and confirmed in squid product. Furthermore, the thermal decomposition kinetics of TMAO-Fe(II) combined with the stimulating components were determined to explore the thermal reaction mechanism of TMAO to DMA and FA. The main findings were as follows:1. An ion chromatography method with non-suppressed conductivity detection was built for the simultaneous determination of DMA, TMA and TMAO. The intrinsic amounts of DMA, TMA and TMAO in marine products were determined by the established method. DMA, TMA and TMAO were separated well by means of ion-exchange chromatography using a5mM nitric acid solution and an IonPac CS17column (250×4mm i.d.) separation column within20mins. Detction limits of DMA, TMA and TMAO were0.030,0.04and0.06mg/L, corresponding to1.01,1.32and2.16mg per kilogram aquatic products, respectively. The relative standard deviations (RSDs, N=6) were between0.83%and2.47%, while the recoveries were between71.62%and103.30%. The content of DMA, TMA and TMAO in four species of marine products including29samples were0-323.55,0-467.41and13.24～10251.39mg/kg. The proposed method has the advantages of high speed, accuracy, sensitivity and accurated properties, which was primarily applicable to the determination of DMA, TMA and TMAO in aquatic products.2. The similar decomposition of TMAO to DMA, FA and TMA was observed in the squid mantle and supernatant during the heating process. Apparently, the thermal conversion of TMAO in squid showed a temperature dependence. Differences of the level of crude protein, crude fat, amino acid (AA), reducing sugar (RS) and reducing activity were determined between raw and boiled squid. The amount of AA (Lys, Glu acid and Met), RS and reducing activity decreased significantly. In addition, the influences of different AA and RS on FA formation in squid supernatant, TMAO and TMAO-Fe(II) aqueous solution were studied at high temperature. The level of FA in the squid extract and TMAO-Fe(Ⅱ) solution increased significantly in the addition of Cys, Lys, Gal and Lev. Cys, Lys, Gal and Lev also promoted the thermal decomposition of TMAO, except Cys only at a lower concentrations (5-30mM). Additionally, radical-(CH3)3N-was detected in squid supernatant and TMAO-Fe(II) by electron spin resonance(ESR). The addition of Lys and Gal enhanced greatly the signal intensity in the squid extract. Therefore, Lys and Gal could promote the conversion of TMAO to FA by enhancing the signal strength of-(CH3)3N-in squid.3. Three model systems TMAO-Fe(II), TMAO-Fe(II)-Lys and TMAO-Fe(Il)-Gal were established, the dicating characteristics and reaction kinetics for the thermal decomposition of TMAO were researched. As the results showed, the thermal decomposition of TMAO was divided into two steps, the former step last5minutes and the latter one last55minutes. The degradation of TMAO was significantly promoted by the addition of Lys and Gal. Zero kinetic rate constants for TMAO, DMA, TMA and FA and the standard error were calculated by linear regression analysis. Temperature and time were the important factors of the thermal decomposition of TMAO, the activation energies of step2were all lower than stepl. The activation energies of FA of stepl and step2in TMAO-Fe(Ⅱ) were68.77kJ/mol and29.72kJ/mol respectively, the sensibility to temperature of step2was lower than stepl. When Gal or Lys was added into TMAO-Fe(Ⅱ), the activation energy of two steps turned lower. The activation energy of FA of stepl in TMAO-Fe(Ⅱ) was68.77kJ/mol, but were25.39kJ/mol and37.66kJ/mol in TMAO-Fe(Ⅱ)-Lys and TMAO-Fe(Ⅱ)-Gal, respectively. Regarding to this datas, lower reaction temperature, less reaction time and the absence of Lys and Gal should be adopted to control the thermal decomposition of TMAO to FA.