Tgp chi Hoa hgc, T. 47 (6), Tr. 753 - 757, 2009
NGHIEN CLTU SlJ PHAN HUY IN VITRO CUA POLYLACTIT
DenTda soan 27-4-2009 MAI VAN TIEN, PHAM THE TRJNH
Vien Hod hgc Cdng nghiep Viet Nam
ABSTRACT
In this study arange of homopoly(lactide) were synthesized hy hulk copolymerization and subjected to in vitro hydrolytic degradation. In vitro degradation studied was conducted at 37"C in pH 7.4 phosphate buffered saline. The effect of lactide type and molecular weight on biodegradable of PEA was studied. The degradation products form in the saline buffer solutions further characterized hy SEM. IR. 'H-NMR; '-C-NMR and GC-MS.
I - DAT VAN DE
Su phat triln ciia ITnh vuc vat lieu sinh hgc trong nhirng nam gin day, dac biet phin Icm cac cdng trinh nghien cii'u tap trung vao cac loai polyme cd khi nang phan hiiy sinh hgc. Da cd rit nhilu san pham dugc phat trien tir polylactic axit (PLA) va cae copolyme ciia chiing nhu poly(laetide-co-glycolide) (PLGA) da dugc irng dung rdng rai trong y tl, nhu chi khau tu tieu, vat lieu ciy ghep md, he giai phdng thudc...[1].
Nhirng sin phim nay ngay cang dugc irng dung rong rai, vi chiing cd tinh tuang hgp sinh hgc cao, cd kha nang phan hiiy bing thiiy phan cac lien kit este td't, sin pham phan hiiy khdng dgc va rit dl gia cong theo nhilu hinh dang khac nhau [2]. Thdi gian phan hiiy in vitro ciia PLA thay ddi trong mdt khoang thdi gian rgng tir 2 - 36 thang. Tdc do va kha nang phan hiiy ciia PLA phu thugc vao nhilu yeu td nhu: nhiet do, do pH, mdi trudng thir nghiem, khdi lugng phan tir va loai polylactic [3]. Trong bai bao nay chiing toi trinh bay mot sd ket qua budc diu nghien ciru sir phan hiiy thiiy phan ciia PLA trong in vitro.
II - THUC NGHIEM 1. Nguyen lieu, hda chait
- Polylactic: Tdng hgp tai VHHCNVN nhu md ta trong tai lieu [5].
- Na2HR04: ham lugng > 997c hang san xua't Merck (Dire).
- NaH2P04: ham lugng > 987c hang sin xuat Merck (Due).
- NaN3: ham lugng > 99,5% hang sin xuat Merck (Diic).
2. Phuong phap nghien cihi su phan huy thiiy phan
a) Chudn bi mdu
De nghien cuu su phan hiiy thiiy phan, cac miu mang da dugc chuan bj nhu sau: tiln hanh hda tan PLA trong dichlorometan de thu dugc dung djch polyme cd ndng do 0,16 g/ml. Sau dd tao mang bing each dd dung dich da pha len dia thiiy tinh rdi cho dung moi bay hai d nhiet do phdng. Sau khoing 5 gid sau khi dung moi da bay hit mang PLA hinh thanh tren dia dugc bdc tich va dem di siy kho a 60"C trong chan khdng sau 24 gid trudc khi tiln hanh nghien ciiu sir phan hiiy thiiy phan trong in vivo. DTa cd dudng kfnh li 20 mm, mdi ming tao thanh cd khdi lugng xip xi 0,5 g.
b) Sif phdn hiiy thiiy phdn ciia PLA trong in vitro
Cac miu chfnh cho nghien ciiu phan hiiy thiiy phan chuin bj trudc dugc dat trong mdt dung djch mudi dem pH = 7,4 tai 37°C. Thanh phin dung dich mudi dem bao gdm: 9 g NaQ, 10,73 g Na2HP04.7H20^ 2,12 g NaH2P04 dugc pha trong 1 1ft nudc cit, pH ciia dung djch dem dugc dilu chinh tdi 7,4 bing each them NaOH.
Cac miu thf nghiem dugc thuc hien trong 10 ml dung djch mudi dem. De ngan chan sir phat trien ciia vi khuan, 1 ml eua dung djch NaN3 ndng do 0,04% da dugc them vao. Cae miu dugc lie diu nhe nhang tren m i y lie ngang trong sudt thdi gian phan hiiy. Sau cac khoing thdi gian nhat djnh la 1, 2, 5, 10, 15 va 23 tuin cac miu dugc la'y ra phan tich. De phan tfch xac djnh cac sin phim phan hiiy, 2 ml dung djch mudi dem da dat miu dugc lay ra phan tfch tren thiit bj GC- MS de xac djnh thanh phin phan hiiy. Mang polyme cdn lai dugc rira bing nudc cat v i siy khd trudc khi phan tfch cac thdng so khac [1,4].
3. Cac phuong phap phan tich dac trung caiu triic tinh chat cua san pham phan huy
- Xac djnh khd'i lugng phan tir ciia polyme bang phucmg phap do do nhdt va phucmg phap sac ky thim qua gel.
- Phuang phap chup inh SEM de xac dinh ciu true bl mat
- GC-MS phan tfch xae djnh sin phim phan hiiy dugc thuc hien tren thie't bj Agilent Technologies 7890A GC-System (Nhat Bin).
- Dd bin keo diit theo deu chuin ISO-527-2 (1993) tren thie't bj do da nang Housfield ciia Anh.
m - KET QUA v A THAO LUAN 1. Dpng hpc phan huy trong in vitro
Sir cd mat cac lidn kit este trong polylactide cho phep sir phan hiiy thiiy phan ciia polyme diin ra theo ca c h i bac. Ca che sir phan hiiy trong in vitro ciia homopolylaetic tdng hgp trong nghien ciiu nay cd thi dugc minh chiimg thdng qua phep do do nhdt dac trung v i su mat khdi lugfng. Hinh la va lb chi ra su suy giam do nhdt dae trung va su mat khdi lugng ciia PLA theo thdi gian thir nghiem. Kit qui hinh la cho thiy do nhdt dac trung ciia polyme giam lien tuc sau khi dat vao mdi trudng dung djch mudi dem d 37°C.
Tir hinh la cho thiy, sir suy giam do nhdt dac trung ciia cic polylactide vdi khd'i lugng phan tir khic nhau dugc the hien vdi td'c do suy giim xip XI nhau v i xay ra ngay sau khi cac miu PLA dugc dat vao dung djch mudi dem.
1.2
61500g/ntol 44000 s PLA:30100 2/i
I g/n ol I a/niol
50 100 150 200
Thai gian [ngay
Hinh la: Su suy giim do nhdt theo thdi gian phan hiiy
500g/riol 44000g/ noi
OOg/inol
50 100 150 200
Thai gian [ngay]
Hinh lb: Sir mit khd'i lugng theo thdi gian phan hiiy Tuy nhien su mat khd'i lugng lai hoan toan
khac, sir mat khd'i lugng chi xiy ra sau mdt khoang thdi gian nhit djnh tiiy thudc vao KEPT ban diu ciia polyme. Su mit khdi lugng tang nhanh khi thdi gian thir tang len dac biet la sau han 120 ngay thir nghiem. Cac polyme cd KEPT thip bj mat khdi lugng nhanh ban so vdi cac polyme cd KEPT cao hem, tai ciing mdt thdi gian phan hiiy.
2. Sir thay ddi khdi lugng phan tu va suy giam dp ben keo cua PLA trong qua trinh phan huy
Cae miu dugc gia cdng theo tieu chuan, dugc ngam trong mdi trudng dung djch mudi
dem. Thf nghiem thue hien cho ba miu PLA cd KEPT ban diu khac nhau de so sanh. Sau 30 ngay djnh ky mdt lin, miu dugc lay ra, lau sach, say khd, dua di xac djnh do bin keo va pha thanh dung djch cd ndng do 0,1% trong chloroform d 25°C dl xac djnh lai KEPT. Kit qua khao sat su thay ddi KEPT, su thay ddi do bin keo ciia PLA dugc trinh bay trong bang 1.
Kit qua trong bang 1 cho thiy KEPT va do bin keo ciia cac miu giam di khi keo dai thdi gian ngam miu trong dung djch mudi dem. Do suy giim KEPT va do bin keo ciia PLA xay ra nhanh ban sau 120 ngay thir. Cae miu PLA cd KEPT cao hem cd thdi gian phan buy dai ban. 0 Bdng 1: Sir thay ddi KEPT va do ben keo theo thdi gian phan hiiy ciia cac miu PLA
Thdi gian phan hiiy, ngiy
0 30 60 90 120 150 180
Miu PLA: 61500g/mol KEPT
61500 57500 30700 21020 15360 8070 2870
5K, MPa 38,1
- 14,7
- - 3,8
-
M i u PLA: 44000g/mol KEPT
44000 36040 24040 15990 9030 3140 630
5K, MPa 35,4
- 13,6
- 5,4
- -
Miu PLA: 30100g/mol KEPT
30100 24260 16060 7870 2600 450
-
5K, MPa 32,2
- 8,6
- 2,2
- -
ciing mdt gii trj KEPT miu PLA da bj thiiy phan cho do bin keo thip ban rit nhilu so vdi miu PLA chua thiiy phan. Nhin chung do bin keo ciia tit c i eac miu PLA sau khi phan hiiy trong mdi trudng in vitro diu giam manh din din nhirng thir nghiem tilp theo la khd khan, tham chf khdng the xac dinh dugc do bin keo trong thdi gian sau dugc nira.
3. Nghien ciifu san pham ciia su phan huy thuy phan
Sin phim phan hiiy hinh thanh trong sud't qua trinh phan hiiy thiiy phan ciia PLA trong dung djch mudi dem photphat dugc xac dinh
bing sic ky khdi phd. Cic hgp chit tao ra tir su phan hiiy cua polyme dugc xac djnh bing cich so sanh vdi thu vien phd va cac miu thir. Hinh 2a va 2b chi ra phd khdi lugng ciia lactic axit va giin dd sic ky khi cua sin phim thiiy phan hinh thanh sau 1,10 va 23 tuin.
Kit qua phan tfch sin pham phan hiiy hinh 2b cho thiy lactic axit, 2-hydroxyl valeric metyl este hinh thanh trong sudt q u i trinh phan hiiy.
Pie sd 1 l i 2-hydroxyl valeric, sd 2 la lactic axit, sd 3 la metyl este. Su nghien cuu v i xac djnh sin phim pMn hiiy l i rit quan trgng, cho phep du bio kha nang k h i nang phan hiiy va tfnh tucmg hgp sinh hgc ciia vat lieu.
I " „ 73
119
138 ISl 1 « ' r i 8 1 " '
30 40 50 60 70
S3 " 67 " Bl ' ^ 1 " ' 138 I S l ^•" j 181 i iLLJli„.,ll,|lll,lilJ|li.i|iim|,l~ili|ilJ |lll|lll,ll|llllpl^^•:•ll|llMlJ^ll'l^l'''il'|l ^"'i'
' - 90 100 110 1!C 130 I M ISO 160 170 180 190 20i
Hinh 2a: Phd khdi lugng ciia lactic axit
"*'tr^|>'i ^ n m iM,..^^
Hinh 2b: Giin dd GC san pham phan hiiy sau 1,10 v i 23 tuin
4. Nghien cim sir thay ddi hinh thai hpc va caiu true be mat mau san pham b i n g phuoTig phap kinh hien vi dien t u quet
Anh SEM tren hinh 3a, 3b,3c, ciu true hinh thai bl mat ciia PLA sau cac khoang thdi gian phan hiiy khac nhau. Hinh 3a li anh chup chup bl mat cua PLA d thdi dilm ban diu chua xay ra sir phan hiiy, hinh 3b, 3c l i inh chup ciu true hinh thai ciia PLA sau 2 va 4 tuin tiln hanh phan hiiy thiiy phan trong mdi truang invitro. Tir cac anh SEM cd thi thay ro sau 2 tuin bl mat PLA da xua't hien cac vit nut gay, be mat bj an mdn dan. Cau triic bl mat bj biln ddi manh sau 4 tuin ngam trong mdi trudng in vitro. Sir diit
gay ciia mang PLA xuit hien nhilu ban va rd ban. Dilu nay chirng minh rang su phan hiiy ciia PEA da xiy ra.
Ill - KET LUAN
Trong mdi trudng in vitro PEA ngay lap tire bj phan hiiy do phin iing thiiy phan din tdi sir suy giam do nhdt dac trung va KEPT. Khdi lugng cua miu bj tdn thit va tfnh chit bl keo ddng thdi bj giim rd ret theo thdi gian thiiy phan. Tdc do phan hiiy xiy ra nhanh han sau khoang 120 ngay ngam miu.
San phim ciia qua trinh phan buy ciia PLA khdng dgc va cd tfnh tuang hgp sinh hgc eao
chiing la nhirng loai axit lactic, axit valeric da dugc xac dinh vi minh chii'ng bing sac ky khdi phd va so sanh vdi phd chuin.
Cau triic hinh thai ciia PLA trudc va sau thdi gian phan buy dugc thi hien rd net bing SEM.
Hinh 3: Anh SEM ciia PLA (a), PLA sau 2 tuan phan hiiy(b), PLA sau 4 tuan phan hiiy
3.
TAI LIEU THAM KHAO
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Taira and T. Kurokawa J. Control. Rel. 88, 229 - 242 (2003).
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Lai. Polymer, 46, 9784 (2005).
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147 (2002).
5. Mai Van Tiln, Pham The' Trinh, Ngd Duy Cudng. Hgi thao vl vat lieu polyme va compozit-Bg Khoa hgc va cdng nghe.
Chucmg trinh nghien ciru phat triln va irng dung cong nghe vat lieu KC-02/06-10, 128 -
132, Ha Ngi (2008).
Lien he: Pham The Trinh
Vien Hda hgc Cdng nghiep
So 2 Pham Ngu Lao, Hoan Kiim, Ha Ndi.
