V ol. 25 N o. 5 2 0 0 4 5 CHEM ICAL JOU RNAL O F CH IN ESE UN IV ERS IT IES 853 857 1, 1, 1, 1, 2, 1 (1., 130023; 2., 130022) N 2 2N 2 2N 2 (N TG),., Savage D EA E2 (D E52) Sephadex G275., (S trep tococcus zooep id em icus J 18),., 1. 16 10 6. ; ; ; O 657 A 025120790 (2004) 0520853205 (H yaluronic acid, HA ), 10 4 10 7 [1 ]. [2 ]. [3 5 ].,, HA HA, HA, HA [6 9 ]. HA. HA [2 ].,, HA. 20 80, HA,, K im [10 ] N T G, Β2, 6 7 ggl. HA,, HA, HA 1. 88 ggl [11 ]. N 2 2N 2 2N 2 (N T G), (S trep tococcus zooep id em icus J18) [12 ], 2. 0 ggl.,,, HA,. 1 1. 1 S trep tococcus zooep id em icus J18 [12 ]. D EA E2 (D E52) SephadexG275, HA, Sigm a,. B ruker IFS66V ; B ruker AV 400 ; J2810 ; Sh im adzu UV 3100 2 ; Ch rist alpha2124. 1. 2 N T G (S trep tococcus zooep id em icus J 18), 50 g : 2003206205. : ( : 2002, 20258247) ( : 20012112 015338). : (1942 ),,,,. E2m ail: ying@public. cc. jl. cn
854 V ol. 25 30 g N ac l 5 g, K 2H PO 4 1. 5 g, KH 2PO 4 0. 5 g, M gso 4 0. 5 g SD S 0. 75 g [12 ]. 250 ml 25 ml, 200 rgm in 37 24 h. 5 000 rgm in 10 m in,, 2. 3,, 10 m in,. 1. 3 HA. [13 ] HA. [14 ], 1. 4 15. 0 m g, 5 ml 6 molgl HC l, 2 h, N aoh ph 7. 0, 50 ml. E lson2m organ [15 ] D 2. 3. 0 m g, 100 ml, [13 ]. 1. 5 13 C NM R 100. 57 M H z, 90 4. 3 Λs, 5 Λs,, 5 000, D 2O, 5%, 0. 1. KB r, B ruker IFS66V, 4 000 400 cm - 1. HA 1 m ggml,. 250 180 nm, 200 nm gm in. 3. 1. 6 2. 5 10-5 molgl, HA 5 m ggml. N aoh (0. 05 0. 4 molgl ). [ 16 ] HA 1 1, N aoh, 15 m in, N aoh ( ), 530 480 nm. 1. 7 HA. [ 17 ], 0. 5 mm, 25, 0. 2 molgl N ac l, HA 0. 2 10-3 0. 5 10-3 ggml, t0 ( ) ti ( ), Γr, Γsp [Γ],. 2 2. 1 2. 1. 1 Savage HA, HA 2. 0 ggl. 1 000 ml 500 ml, Savage [18 ]. 500 ml g [V ( ) V ( ) = 4 1 ],, 4 000 rgm in 20 m in,,, 3,. 2. 3,. 5 000 rgm in 10 m in,, P 2O 5, 1. 14 g. 2. 1. 2 500 ml, 4% (CPC ),. 1 h,. 5 000 rgm in 10 m in, 100 ml 1. 5 molgl N ac l,. 2. 3, P 2O 5 1. 05 g. 2. 1. 3 D EA E2 100 ml, D EA E2 (D E52) (2. 5 cm 20 cm ). 0. 5 3. 0 molgl N ac l, 1 ml gm in, ( 5 ml ),, 1.,,
N o. 5 : 855, 0. 75 g. F ig. 1 Elution curve of polysacchar ide on F ig. 2 Elution curve of polysacchar ide af ter gel D EAE-cellulose column f iltra tion on Sephadex G-75 column 2. 1. 4 Sephadex G275 0. 5%, Sephadex G2 75 (1. 5 cm 120 cm ), 2. 0 ml,, 6 ml gh,, 2., 0. 68 g. Savage D EA E2 SephadexG275, 59. 65%, 0. 075%. 2. 2 2. 2. 1. 30 ΛggmL, 13. 74 ΛggmL, w 1 (% ) = [ (c1 V 1) gm ] 100% (w 1 ; c1 ; V 1 ; m ), 45. 8%. E lson2m organ N 2. 30 ΛggmL, (10 ΛggmL ) 525 nm, w 2 (% ) = [ (A c1) g(a 0. 830 9 c2) ] 100% (w 2 ; A ; c2 ; A ; c1 ; 0. 830 9 ) 45. 9%. N 1gN 2 = [ (w 1gM r 1 ) g(w 2g M r 2 ) ] (N 1gN 2 ; M r 1 ; M r 2 ), 1 1. 08, HA. 2. 2. 2, 3. COO 173. 70, COO ; O (O )CH 2 102. 71 100. 14 ; > CHOH 5 68. 00 82. 09 ; > CHN H 53. 85 ; CH 2OH 60. 25, 22. 03 CH 3. 13 C NM R,. F ig. 3 13 C NM R spectrum of the pur if ied polysacchar ide F ig. 4 IR spectra of the pur if ied polysacchar ide(a) and standard HA (b) 2. 2. 3 4. 4
856 V ol. 25 3 400 cm - 1 OH ; 2 940 cm - 1, CH 2 CH 3 C H ; 1 560 1 620 cm - 1 C O C N, ; 1 400 1 320 cm - 1 COO C O OH 2,. HA. 2. 2. 4, Cotton. ( ) ( ), 230 nm. Χ2Π ( 208 212 nm ), Cotton, Cotton Π2 Π 3. O 24 Π2Π 3 Cotton, O 23 Π2Π 3 Cotton [19 ]. HA, 5., 209. 57 nm Cotton,. 184. 32 nm Cotton, O 23, HA,. F ig. 5 C ircular dichro ic spectra of the pur if ied HA and standard HA a. Purified polysaccharide; b. standard HA. F ig. 6 Change in the absorption max ium of the Congo red-ha com plex a. Congo red2ha comp lex; b. Congo red. 2. 2. 5 (Congo red), C 32H 22N 6O 6S2N a2,, N aoh,. [20 ]. HA 6,,,. HA COO -,,. [21 ], HA. HA,. 2. 2. 6 HA HA [Γ]= 19. 74, [Γ]= 3. 6 10-4 M 0. 78 r HA 1. 16 10 6.,, HA,, 1. 16 10 6. [ 1 ] L ap ik L. J ṙ, L ap ik L., De Sm edt S. et al.. Chem. Rev. [J ], 1998, 98 (8) : 2 663 2 684 [ 2 ] ZHOU Rong2Q ing ( ), L IL in ( ), GUO Si2Yuan ( ). Sugarcane and Canesugar ( ) [J ], 2001, (6) : 51 57 [ 3 ] A kim a K., Ito H., Iw ata Y. et al.. J. D rug Target[J ], 1996, 4 (1) : 1 8 [ 4 ] M uckenschnabel İ, Bernhardt G., Sp russ T. et al.. Cancer Chemotheṙ Pharm acol. [J ], 1996: 38 (1) : 88 94
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