The Extraction Of Flavor Compounds From Wasabi And Their Antibacterial Activity

Wasabi, which is a kind of cruciferous plants, its main flavor component is isothiocyanates. Isothiocyanates do not present in the wasabi directly. When the cells of wasabi are damaged under the external action, glucosinolates in cell vacuoles are enzymatically hydrolyzed by myrosinase. Different R side chain groups form different isothiocyanates, and the main component of isothiocyanates is allyl isothiocyanate (> 80%). Isothiocyanates has a special pungent taste, they can not only be eaten, but also have impact on inhibition of bacterium, cancers and so on. Wasabi is widely studied and applied in Japan and South Korea, however, researches on wasabi are relatively less in our country, and most studies are focus on isothiocyanates from horseradish and mustard. In this study, quantitative analysis method of isothiocyanate from wasabi is established, extraction methods of isothiocyanates from wasabi are explored and antibacterial activity of wasabi extraction is tested. The main contents and results are as follows:Established quantitative analysis method of isothiocyanate from wasabi by ultraviolet spectrophotometry (UV) and gas chromatography (GC). In the analysis of UV, isothiocyanates have maximum absorption at the wavelength of 243nm. Through experiments, the standard curve of isothiocyanates in wasabi by UV is obtained: y=8.3488x+0.09317, and correlation coefficient is 0.9992, ranging in 0.05050mg/mL-0.3030 mg/mL. Recovery rates are 99.84%-106.9%、97.20%-104.1%、97.64%-101.2% respectively in different amount of standard added, RSDs are 2.74%、2.42%、1.43% respectively. The recovery rate is high, the quantitative results are accurate and reliable. What’s more, this method has the advantages of simple operation and short experimental time. It is a fast, convenient method for quantifying total isothiocyanates.Quantitative analysis method of isothiocyanate from wasabi by internal standard method of GC is also established. Agilent 7890A gas chromatography with FID detector is used. The operating conditions for GC were as follows:injector and detector temperature, 250℃; HP-5 (30M×0.32mm) column. column temperature 50℃ for 5 min, then increasing to 100℃ at 5℃/min, keep 100℃ for 2 min, then increasing to 250℃ at 15℃/min, keep 250。C for 2 min. The split ratio is 1:20. Butyl acetate is used as internal standard. Through experiments, the standard curve of isothiocyanates in wasabi by GC is obtained: y = L3166x+ 0.0079,and correlation coefficient is 0.9999,ranging in 0.5095 mg/mL-lO.Olmg/mL. The RSD of the precision test of the method is 0.12%; The RSD of the stability test is 0.24%;Recovery rates are 99.56%-106,2%%, 97.21%-101.5%, 97.29%%-100.2% respectively in different amount of standard added, RSDs are 2.32%, 1.62%, 1.14% respectively. It shows that the method has high precision, good stability and high recovery rate. It can not only determine the content of allyl isothiocyanate, but also can determine multiple isothiocyanates.Single factor experiment and orthogonal optimization experiment are used to explore the extraction of wasabi in isothiocyanates by ultrasound-assisted extraction. On the basis of single factor experiments, using five factors: hydrolysis temperature, hydrolysis time, pH,ultrasonic time and ultrasonic power,four levels of each factor(5’ to design orthogonal experiment. The results showed that the order of five factors affecting the extraction rate is:hydrolysis temperature > pH > hydrolysis time > ultrasonic power > ultrasonic time. The optimal combination is that hydrolyzing for Ih at 40。C, pH is 6,and ultrasonic testing for lOmin at the power of 40 W. We can get the highest extraction rate of isothiocyanates in this condition of 0.7829 mg/g.In addition,the feasibility of extracting isothiocyanates from wasabi by supercritical carbon dioxide extraction is studied. The extraction temperature is 40。C and extraction pressure is 20 MPa. The products are characterized by GC. Results show that we can extract isothiocyanates by supercritical carbon dioxide extraction. Results show that using 10%ethanol as cosolvent,can increase the extraction rate. Experiments illustrate that extracting isothiocyanates from wasabi by supercritical carbon dioxide extraction is feasible.Study the antibacterial activity of wasabi extraction on common bacterium(Staphylococcus aureus,Candida albicans,Bacillus cereus Frankland, Pseudomonas aeruginosa, Vibrio Parahaemolyticus, Escherichia coli,Proteusbacillus vulgaris) and zoonotic bacterium(Streptococcus suis, Streptococcus agalactiae, Erysipelothrix rhusiopathiae, Bordetella bronchiseptica, Riemerella anatipestifer) by inhibition zone test and minimum inhibitory concentration test. Results show that wasabi extraction have inhibitory effect on the 12 kinds of bacterium, and the antibacterial activity of wasabi extraction is stronger than standard allyl isothiocyanate. What’s more, the inhibition of wasabi extraction on the gram positive bacteria is stronger than on gram negative bacteria and zoonotic bacterium also accord with the laws.