cus sinensis 3 50 (2) : , 2004 ZHOU Xian2Qing 1,2, XIE Meng2Xia 3, N IU Cui2J uan 1 33, SUN Ru2Yong 1 ν 2004 Acta Zoologica Sinica

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1 50 (2) : , 2004 A cta Zoologica S inica C E C E 3 1, , , , C ( ) E (V E ) V E, 5, V E 0 0 mg/ kg ( ) mg/ kg () mg/ kg () ; mg/ kg ( ) ; mg/ kg ( ) V E - V E, - V E V E, - V E, V E, ; - V E, 4 -, -,,, 4, -, -, V E,, V E [ 50 (2) : , 2004 ] C E Effects of the combination of vitamin C and E on growth, liver vitamin C and E, and levels of serum cortisol in stressed and non2stressed juvenile sof t2shelled turtles Pelodis2 cus sinensis 3 ZHOU Xian2Qing 1,2, XIE Meng2Xia 3, N IU Cui2J uan 1 33, SUN Ru2Yong 1 11 College of Life Sciences, Beijing Normal University, Ministry of Education Key Laboratory for Biodiversity Science and Ecological En2 gineering, Beijing , China 21Department of Laboratory Animal Science, Capital University of Medical Sciences, Beijing , China 31Centre of Laboratory, Beijing Normal University, Beijing , China Abstract We studied the effects of the combination of vitamin C and E on growth, liver vitamin C and E, and level of serum cortisol in stressed and non2stressed juvenile soft2shelled turtles. The turtles were fed combination of vitamin C and E at the dosages of 0 and 0 mg/ kg diets (control group), 250 and 50 mg/ kg diets (experiment group ), and 50 mg/ kg diets (experiment group ), 250 and 250 mg/ kg diets (experiment group ), and 250 mg/ kg diets (experiment group ) diets, respectively for 4 weeks. The growth of juvenile soft2shelled turtle was estimated by specif2 ic growth rate, the level of liver vitamin C and E was analyzed by the method of high pressure liquid chromatogram , (No ) [ This research was funded by the National Natural Science Fundation of China (No ) ] 33 (Corresponding author). E2mail : edu. cn ν 2004 Acta Zoologica Sinica

2 2 : C E C E 159 ( HPLC), and serum cortisol was determined by radioimmunoassay. The specific growth rate in the four groups of experi2 ment - were higher than that of the control, whereas no significant difference was observed among experiment - groups. Levels of liver vitamin C and E in non2stressed turtles increased with increase of dietary vitamin C and E, and experiment - groups were significantly higher than those of the control group and experiment group. After acid stress, levels of liver vitamin C and E in all groups had a tendency to decrease, but there were no significant changes ; lev2 els of liver vitamin C and E in experiment - groups were higher than those of the control group, and experiment group was higher than the other four groups after acid stress. The level of serum cortisol showed no significant difference among experiment - groups, and though experiment - groups had a decrease tendency compared with control group, there was no significant change ; whereas experiment groups was lower than that of control. After acid stress, serum cortisol in control group was significantly elevated, whereas the other four groups showed no significant change, al2 though there was a tendency to increase. Serum cortisol levels in experiment - groups were lower than those of the control group after stress, whereas there was no significant difference among - groups. These results suggest that the absence of vitamin C and E depressed growth, and their combination had no interactive effect on the growth and serum cortisol levels, whereas they had an effect on levels of liver vitamin C and E in soft2shelled turtles [ Acta Zoologica Sinica 50 (2) : , 2004 ]. Key words Soft2shelled turtles, Pelodiscus sinensis, Vitamin C, Vitamin E, Acid stress, Growth, Liver, Cortisol C ( ) E (V E ),, 2 h,,, E ( 30 1, 2003a), C, C3 C4 (, V E, 2003b) (1998) 3, V E (, 1996) ( Tanaka et al., 1997) 1 C E V E (Bendich et al., 1984), Table 1 The supplementation of vitamin C and E in diet, V E (, 1996) V E Grouping,, (, 1996), V E, ( Hamilton et al., 2000) Cadenas et al. (1997), V E, DNA,,, V E, V E 112 ( Pelodiscus sinensis) V E 60 (, V E g), 5 12, 60 cm 30 cm 40 cm V E 1, - 20, 4 h 30 1, 6 00,, 4,, V E V E (mg/ kg ) (mg/ kg diet) Control group (mg/ kg ) (mg/ kg diet), 5 V E, 5

3 , - 20 ; t P < 0105,, p H 719 p H 510, 24 h ( 011 g), - : ( Specific growth rate : %/ d) = (lnw 2 - lnw 1 ) / T 100 % T = ( T 2 - T 1 ), T 1 ( F 1,9 = 0137, P = 01576) ( 1), T 2 ; T 1 W 1, T 2 W ,, 20 ml 3 %,, :, KON TRON C 18 (5 m, 416 mm 250 mm) ;, = 6 94 ( v/ v), p H = 2163 ; 243 nm ; 1 ml/ min ; 20 l, ( ) V E C : : 1 (1994),, < 0105) 30 ml, 10 ml 12 %, 10 ml 40 % Fig11 Effects of the combination of vitamin C and E on specif ic growth rate in juvenile soft2shelled turtles ( Mean,, min,, 60 ml 2 1 2, 150 ml, (50 ml 3), 0105). V E ( P < 0105), - ( P > 0105), V E 1 C E Duncan, ( P SD) C: Control group. : Same as those in the Table 1. Duncan multiple range test. The values with different superscript letter are significant different among different dose groups ( P <, 212 V, 2 ml,, C V : KON2 C V E TRON C 18 (5 m, 416 mm 250 mm) ;,, ( F 1,24 = 41456, P = 97 3 (v/ v) ; 290 nm ; 1 = 01048) ml/ min ; 20 l, ( F 1, 24 = , P = ),, V E ( F 1,24 = , P = 0101) Duncan, ( P < 0101) ( 2) 213 V E STA TISTICA V E,, V E V E ( F 1, 24 = , P = ), Duncan ( F 1, 24 = , P = 01001),

4 2 : C E C E C E C ( ) Duncan : ( P < 0105) ; t : P Fig12 Effects of the combination of vitamin C and E on content of liver vitamin C in acid2stressed and unstressed juvenile soft2shelled turtles ( Mean + SD) Duncan multiple range test : different superscript letter are significant different among different dose groups in unstressed soft2shelled turtles ( P < 0105). t2test : P values is the result of comparison before and after acid2stress for per dose groups. 3 C E E ( ) Duncan : ( P < 0105) ; t : P Fig13 Effects of the combination of vitamin C and E on content of liver vitamin E in acid2stressed and unstressed juvenile soft2shelled turtles ( Mean + SD) Duncan multiple range test : The values with different superscript letter are significant different among different dose groups in unstressed soft2shelled turtles ( P < 0105). t2test : P values is the result of comparison before and after acid2 stress for per dose groups.

5 V E ( F 1,24 = ( F 1,26 = 0, P = 1100), , P = 01045) - V E - ( P < 0101) ( 3) ( P > 0105), 214 ( P < 0101) ( 4) V E 4 C E ( ) Duncan : ( P < 0105) ; t : P Fig14 Effects of the combination of vitamin C and E on content of serum cortisol in acid2stressed and unstressed juvenile soft2shelled turtles ( Mean + SD) Duncan multiple range test : different superscript letter are significant different among different dose groups in unstressed soft2shelled turtles ( P < 0105). groups. t2test : P values is the result of comparison before and after acid2stress for per dose 215 V E ( F 1,26 = V E V E, ( P > 311 V E 0105) - V E ( P < 0101),, V E 4 ( P < 0101) V E (1996) V C ( F 1,24 = 41874, P = ) V E Tanaka et al. (1997) ( F 1,23 = 41917, P = 01039), V E ( 2, 3) V E,, ( P = 01004), 4, 312 V E ( P > 0105) - ( P < 0105), - ( P > 0105) V E, 01069, P = 018) ( 4) 3, V E V E, V E Bendich et al.

6 2 : C E C E 163 (1984), 012 g/ kg, V E, 314 V E V E 10 g/ kg, V E ( mg/ kg) V E, V E (, 1996) ; 10 3 h, ( Hamilton et al., (, 1997) ; 2000) V E 3 V E (,, V 1992), V E C V E (, 1995),, V E, ( Tanaka et al., 1997), V E V E V E,,, V E 315 V E, V E,, V V E ( E - ),, V E, ; V E,,,, ( ), ( Frei et al., 1989) ( Sunyer et al., 313 V E V E 1995 ;, 1997),,, V E, V E, V (10 g/ kg) V E (Ben2 E, V dich et al., 1984) C V E, ( Kanazawa et al., 1981 ; Hruba et al., 1982), V ( C V E Tanaka, V E et al., 1997) V E V E, V E (, 1996), V E, V E V E,,, V E V E,,, V E V E : 1) V E, V E ; ( References) ( ) (Niki et al., 1984) (Doba et al., 1985) ( Wefers and Sies, 1988 ; Mehlhorn et al., 1989) V E V E, ; 2) V E, V E Bendich A, Dapolito P, Gabriel E, Machlin LJ, Interaction of dietary vitamin C and vitamin E on guinea pig immune responses to mitogens. J. Nutr. 114 : Cadenas S, Barja G, Poulsen HE, Loft S, Oxidative DNA damage estimated by oxo8dg in the liver of guinea2pigs supple2 mented with graded dietary doses of ascorbic acid and alpha2toco2 pherol. Carcinogenesis 18 (12) :

7 Doba T, Burton GW, Ingold KU, Antioxidant and co2antioxi2 dant activity of vitamin C : the effect of vitamin C, either alone or in the present of vitamin E or a water soluble vitamin E analogue, upon the peroxidation of aqueous multilamellar phoshohpid lipo2 somes. Biochim. Biophs. Acta 835 : Frei B, England L, Ames BN, Ascorbate is an outstanding an2 tioxidant in human blood plasma. Proc. Natl. Acad. Sci. USA 86 : Gao L X, Xu D Y, Jin H, A study on the adequate dosage of vita2 min E in prevention and treatment of visceral injury in burned rats. Acta Nutrimenta Sinica 14 (4), ( In Chinese). Gao L X, Jin H, Xu ZQ, Wang ZY, Xu D Y, Gu J F, The metabolic changes of vitamin A, E, C and adequate supplementa2 tion after burn and trauma. Acta Nutrimenta Sinica 17 (3) : ( In Chinese). Hamilton IM, Gilmore WS, Benzie IF, Mulholland CW, Strain JJ, Interactions between vitamins C and E in human subjects. Br. J. Nutr. 84 (3) : Hruba F, Novakova V, Ginter E, The effect of chronic marginal vitamin C defeiciency on the 2tocopherol content of the organs and plasma of guinea2pigs. Experientia 38 : Kanazawa K, Takeuchi S, Hasegawa R, Influence of ascorbic acid deficiency on the level of non2protein SH compounds and vita2 min E in the blood and tissues of guinea pigs. Nihon. Univ. J. Med. 23 : Li AH, The effects of crowding on plasma cortisol, blood sugar and liver ascorbic acid content in grass carp. Acta Hydrobiological Sinica 21 (4) : ( In Chinese). Mehlhorn RJ, Sumida S, Packer L, Tocopheroxyl radical persis2 tence and tocopherol consumption in liposomes and in vitamin E2en2 riched rat liver mitochondria and microsomes. J. Biol. Chem. 264 : Niki E, Saito T, Kawakami A, Kamiya Y, Inhibition of oxida2 tion of methyl linoleate in solution by vitamin E and vitamin C. J. Biol. Chem. 259 (7) : Sunyer JO, Gomez E, Navarro V, Quesada J, Tort L, Physio2 logical responses and depression of humoral components of the im2 mune system in gilthead sea bream Sparus aurata following daily acute stress. Canadian Journal of Fisheries and Aquatic Sciences 52 (11) : Tanaka K, Hashimoto T, Tokumaru S, Iguchi H, Kojo S, In2 teractions between vitamin C and vitamin E are observed in tissues of inherently scorbutic rats. J. Nutr. 127 : Wefers H, Sies H, The protection by ascorbate and glutathione against microsomal lipid peroxidation is dependent on vitamin E. Eur. J. Biochem. 174 : Wen J, Lin J H, Wang HM, The effects of dietary vitamin E and ascorbic acid on growth and immunity in chicken. Acta Verinariaet Zootechnica Sinica 27 (6) : ( In Chinese). Xu L H, Chen Z, Xu Y, Zhang YY, The method of high perfor2 mance liquid phase chromatography determining vitamin E in fish. Acta Hydrobiologica Sinica 18 (2) : ( In Chinese). Zhang FM, Zhang SQ, Wang XF, Effects of and V E on ac2 tivity of myocardial cells from virus infection. Acta Nutrimenta Sinica 18 (4) : ( In Chinese). Zhou XQ, Niu CJ, Li QF, The effects of light intensity on daily food consumption and specific growth rate of the juvenile soft2 shelled turtles Trionyx sinensis. Acta Zool. Sin. 44 (2) : ( In Chinese). Zhou XQ, Niu CJ, Sun RY, 2003a. The effects of vitamin E on growth, liver vitamin E and serum cortisol level in juvenile soft2 shelled turtles Pelodiscus sinensis. Acta Zool. Sin. 49 (1) : ( In Chinese). Zhou XQ, Niu CJ, Sun R Y, 2003b. Effects of vitamin C and acid2 stress on serum complement C3 and C4 level in juvenile soft2shelled turtles Pelodiscus sinensis. Acta Zool. Sin. 49 ( 6) : ( In Chinese).,,, E. 14 (4) : ,,,,,, A E C. 17 (3) : , (4) : ,,, E. 27 (6) : ,,,, E. 18 (2) : ,,, C E. 18 (4) : ,,, (2) : ,,, 2003a. E E. 49 (1) : ,,, 2003b. C C3 C4. 49 (6) :

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, ) ( mg/ kg 4, C3

, ) ( mg/ kg 4, C3 49 (6) :769 774, 2003 A cta Zoologica S inica C 3 C3 C4 33 (, 100875) (, 100054) C ( Pelodiscus sinensis) C3 C4, 6, C 0 250 500 2 500 5 000 10 000 mg/ kg, 4, C3 C4, C 250 mg/ kg, C3 ; C 500 2 500 5 000