37 4 2007 4 PERIODICAL OF OCEAN UNIVERSITY OF CHINA 37 (4) :586 590 J uly, 2007 Ξ, ΞΞ, (, 266003) :, ( A postichopus japonicus) (LDH) ( EST) (SOD) LDH 10 20,12 h EST SOD, 20 10 1 h EST 3 h SOD,EST SOD 1,, : ; ; ; : S917. 4 : A : 167225174 (2007) 042586205, [1 ] [225 ] (LDH), [5 ], [6 ],LDH, LDH [7 ] ( EST) [8 ] (SOD), [9210 ],SOD, SOD [11 ] ( A postichopus japonicus), [12213 ],, [14215 ],,,, 1 ( 2. 5 g) 10 20 2, ; 18 2 1. 2 1. 2. 1 2, 10 20 0 h,1 h,3 h,12 h,24 h,72 h 10, - 80, 20 10 1. 2. 2 18, (18 2), (18 4), (18 6) ( 1), 18 45 d, 10, - 80 1. 1 1 1 Fig. 1 Diagram of the diel temperature fluctuating mode Ξ ΞΞ : (30400333) :2006210216 ; :2007201205 : (19812),,, E2mail :pinkhum - cn @sina. com : E2mail : dongyw @ouc. edu. cn
4, : 587 1. 3,, ( 0. 05 mol/ L,pH = 75) 4 3 000 g 5 min, -, - 20 1. 4 3 10 % 4. 5 % Tris2HCl (0. 5 mol/ L,p H = 6. 8) Tris2HCl (1. 5 mol/ L, p H = 8. 8) Tris2 (p H = 8. 3) 4, 20 ma, 40 ma, 1 cm [16 ] : NAD 40 mg, 0. 925 ml,pms 2 mg,nb T 20 mg,0. 5 mol/ L (p H = 7. 5) 10 ml,0. 1 mol/ L NaCl 5 ml, 50 ml [17 ] : 50 mg 2 50 mg 2, 50 mg,, 50 ml 0. 1 mol/ L (ph = 7. 0), 37,, 7 % [18 ] : 2. 45 10-3 mol/ L (NB T) 20 min, 0. 028 mol/ L ( TEMED) 2. 8 10-5 mol/ L 0. 030 mol/ L (p H = 7. 8) 15 min, 0. 05 mol/ L (ph = 7. 8) 10-4 mol/ L EDTA, 4 000 lx 30 min, [17 ], 1, EST1, 20 10,1 h EST2, 3 h EST2, ( 2) 2 3 Fig. 2 Electrophoretograms of EST isozyme at acute temperature changes, (showed the deep color band, Level 1 band) ;, (showed the light color band, Level 2 band) ;, ; (showed the lightest color band, Level 3 band ; the band at every levels di2 vided into the brand band and the narrow band, according to the band width) SOD S21, 10 20, 1 h S22 S23,12 h S22 24 h 20 10 1 h 3 h S22 S23 12 h ( 3) 3 SOD 3 Fig. 3 Electrophoretograms of SOD isozyme at acute temperature changes 2 2, LDH, EST SOD 2. 1 EST SOD EST EST1 10 20,EST2,12 h EST2, 24 h EST2, 4 33 Fig. 4 Electrophoretograms of EST isozyme at different amplitudes of temperature change 3 1 6 10 20 0 h,1 h,3 h,12 h,24 h,72 h 7 12 20 10 0 h 1 h 3 h 12 h 24 h 72 h Lanes 1 6, electrophoretograms at 0 h, 1 h, 3 h, 12 h, 24 h and 72 h heat exposure (from 10 to 20 ), respectively ; Lanes 7 12, electrophoretograms at 0 h, 1 h, 3 h, 12 h, 24 h and 72 h 33 2 5 ( Fig. 2 Fig. 5) 1 2 (18 0) 3 4 (18 2) 5 6 (18 4) 7 8 (18 6)
588 2 0 0 7 2. 2 EST SOD, EST SOD 1, ( 4, 5) 5 33 Fig. 5 Electrophoretograms of SOD isozyme at different 3 amplitudes of temperature change,, [19220 ], [21 ], [22 ], [20 ],, [23 ],,, 1 [24 ], 21 35 6 h,, 12 h 35 21 24 h,,,,,est2 12 h,,, EST2 1 h,, [14 ] [12 ],, [12 ], (reactive oxygen species, ROS) [25226 ],, SOD,SOD, [27 ] [28 ] [29 ] SOD,, [30 ], SOD 10 20 [31 ], (ROS) [32 ] ROS [33 ], SOD SOD,,,, ROS, [34 ], ROS, [26 ] [35 ], ( SOD) [36 ], EST SOD 3, 70 d, 18,(18 2),, [15 ] 45 d,2, EST SOD,,,EST SOD,, 25 g [37 ], [12 ],LDH,,, LDH [38 ]
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590 2 0 0 7 1995, 112C: 3092313. [35 ] Gutteridge J M, Halliwell B. The measurement and mechanism of lipid peroxidation in biological systems [ J ]. Trends Biochem Sci, 1990, 15 (4) : 1292135. [36 ] Parihar M S, Dubey A K, Faveri T, et al. Changes in lipid peroxidation, superoxide dismutase activity, ascorbic acid and phospholipids content in liver of freshwater catfish Heteropneustes Fossilis exposed to elevated tem2 perature [J ]. J therm Biol, 1996, 21 : 3232330. [37 ],,,. ( A postichopus japonicus Selenka) 1 [J ]., 1996, 3 (2) : 41248. [38 ]. [D]. :, 2005. Effects of Temperature Changes on Isozymes in the Sea Cucumber, Apostichopus japonicus J I Ting2Ting, DON G Yun2Wei, DON G Shuang2Lin ( Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, China) Abstract : The effects of acute temperature change and diel fluctuating temperature on the isozyme patterns of LDH, EST and SOD of t he sea cucumber A postichopus japonicus were st udied. temperature change, young A. In t he experiment of acute japonicus were subjected to heat exposure (from 10 to 20 ) and cold expo2 sure (from 20 to 10 ), respectively. The samples were taken at 0 h, 1 h, 3 h, 12 h, 24 h and 72 h after heat or cold exposure, respectively. In t he experiment of diel fluct uating temperat ures, young A. japonicus were subjected to four temperature treatments ( 18, 18 2, 18 4 and 18 6 ). The results showed that there was no expression of LDH isozyme. In the experiment of acute temperature changes, within 12 h heat exposure, there were new isomers that appeared both in EST and SOD. After 1 h cold exposure, there was one new band EST2 in the EST isozyme pattern. After 3 h cold exposure there were two new bands band S2 and band S3 in the SOD isozyme pattern. were no changes in t he patterns of EST and SOD isozymes. In the experiment of diel fluctuating temperature, there Key words : A postichopus japonicus ; temperature change ; isozyme ; LDH ; EST ; SOD