Luan An Tot Nghiep

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Phan 1:

M AU

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Cay ieu (cay ao lon hot) la loai cay nhiet i, de trong, a c trong Viet Nam t lau. Luc au no c trong le te cac a phng va oi nui e chong xoi mon at, thng lay qua an ti. Nhng sau o nhan hat ieu co the xuat khau nen cay ieu ngay cang co gia tr. Viet Nam mi bat au quan tam phat trien trong ieu vi qui mo ln t au nhng nam 80, en nay a tao c 125.000 ha va san lng hat ieu at en 80.000 tan, ng th 3 sau An o va Braxin. Trong ieu lay hat la chu yeu, nhng hat ieu ch chiem 15% cua toan bo trai ieu. Nh vay khi nhan c 80.000 tan hat ieu th cung nhan khoang 900.000 tan trai ieu sau khi lay hat. Phan trai ieu nay cha nhieu ng (khoang 10%), nc (75% 85%), protid (0.9%), acid (0.7%), glucid (6.4%), xenlulose (0.4%), vitamin C (200mg%). Ngoai ra con mot so thanh phan dinh dng khac. Nhng trong trai ieu co cha nhieu tannin. Chnh hp chat nay gay ra v chat cho dch trai ieu va o la tr ngai ln khi s dung trai ieu che bien thc pham. Cho nen phan ln trai ieu c bo i sau khi lay hat. Nh vay chung ta a bo i mot lng ng va vitamin C ang ke. Mat khac viec o ba bai trai ieu co the gay o nhiem moi trng. Nhiem vu cua chung em la tm cach thch hp e tach tannin ra khoi dch chiet trai ieu. Viec tach tannin ra khoi trai ieu kha phc tap. a co nhieu nha nghien cu lien quan en van e nay. Nhng cha at c ket qua nh mong muon. V the en nay, lng trai ieu sau khi thu hoach hat c tan dung che bien ca trong nc va nc ngoai ch mi at mot t le kha thap. Vi mong muon chung la gop phan hoan thien them cac cong trnh nghien cu ve tannin thc vat a c biet t lau, mat khac, gop phan tm bien phap che bien, lam tang gia tr s dung cua trai ieu, em xin a vao e tai : Nghien cu tm mot phng phap trch, lam sach nhom polyphenol trong nc trai ieu Song do phng tien th nghiem con han che nen ket qua ghi nhan c trong luan van nay

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chac chan khong tranh khoi nhng thieu sot. Knh mong quy thay co va oc gia ch dan them e luan van nay c hoan chnh hn.

Luan An Tot Nghiep

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Phan 2:

TONG QUAN

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I. VAI NET VE CAY IEU VA DCH TRAI IEU: 1. S lc ve cay ieu: Cay ieu co ten khoa hoc la Anacardium Ocidental Linne thuoc giong Anacardium, ho Anacadiaceae, bo Rutales. Ho nay gom co 56 giong va khoang 500 loai, vo cay co cha nha. moi nc cay ieu co ten goi khac nhau: Cashew (Anh), Cajou, Acajou, Anacadier (Phap), Cajoucu (Braxin), Kaju (An o), Kajus (Malaixia), Yakoi, Ya-ruang (Thai Lan), Kasry, Kasui, Kasol (Philipine), Gian - hu, Yiaokuo (Trung Quoc)... va Viet Nam goi la cay ieu hay cay ao lon hot. Ve nguon goc, cay ieu la loai cay nhiet i, song hoang dai, cay ieu thng moc cac bai cat ven bien va trong cac rng t nhien quan ao An-ti va Braxin, lu vc song Amazon thuoc Nam My. Cay ieu c trong trc tien e chong xoi mon at cac vung oi nui, trung du va e lay hat, la cay song lau nam. Bnh thng sau 3 nam la cay ieu cho thu hoach. Cay ieu tro bong t thang mot en thang t. S thu phan cua bong nh con trung. Sau khi thu phan, hat ieu hnh thanh va phat trien rat nhanh. Thc chat hat chnh la trai, en mot o ln nhat nh th cuong mi phat trien phnh len thanh trai gia giong nh trai le. Sau khoang 60 ngay th trai chn. Khi chn, trai co mau vang hoac mau o tuy loai. Trai chn co ac iem la vo mong, da lang bong, ben trong cha tht mem, nhieu nc, co v ngot, chua, chat, co mui thm ngat va luc o hat a gia. Hat ieu t khi mi hnh thanh en luc gia co hnh dang cua mot qua than. Luc non co mau xanh la cay sang nhat, luc ve gia co mau xanh nau. La ieu thuoc la n nguyen, moc so le, hnh oval. Hoa th moc thanh chum, mau vang/ trang co gan o hay hong tuy giong. Cay ieu thng co chieu cao t 6m 10m, than ngan, khuc khuyu, am choi rat nhieu, phu canh quanh than chnh va sa xuong tan mat at. Da vao mau sac cua trai ieu ngi ta phan biet thanh hai giong ieu chnh : ieu o va ieu vang. ieu o : co hai loai Loai trai tron nho co v chat manh. 5

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Loai trai dai nhon t, co nhieu nc, v chat t hn va co v ngot cao. ieu vang : co hai loai. Loai trai to, dai, co nhieu nc, v ngot. Loai trai co mau vang nhat (con goi la ieu nep), trai ln, da lang, nhieu nc, v ngot du. V chat la v ac trng va la nguyen nhan han che viec s dung, che bien nc trai ieu. Cay ieu khong chu c ngap ung, a sang va nhiet o thch hp cho cay phat trien la 25 300C. Tuoi tho co ch cua cay ieu keo dai vai chuc nam. San pham chnh cua cay ieu hien nay la hat ieu. Nhan hat ieu dung trong thc pham. Co the khai thac dau trong vo hat e dung trong cong nghiep. Phan trai ieu co the s dung che bien thanh nc giai khat co hoac khong co con, an song nh rau hoac dung lam thc an gia suc. Tuy nhien do co cha nhieu tannin nen pham vi s dung rat han che. Ve nguon goc, theo nhieu tai lieu nghien cu ve cay ieu cho biet: cay ieu a moc hoang dai thanh rng t nhien pha bac Braxin. T nhng nam 1500, ngi Bo ao Nha a du nhap giong ieu en An o nham muc ch trc tien la chong xoi mon at tieu bang Goa. Roi sau o c trong sang nhieu tieu bang khac. Hien nay, cay ieu c tm thay cac nc chau Phi, chau My La tinh, chau A va chau Uc. An o la nc au tien xem cay ieu la hoa mau co gia tr thng mai. Dien tch trong ieu tren the gii ngay cang tang do nhu cau ve hat ieu ngay cang tang. Ngoai s phat trien ve lng, ngi ta con quan tam ve chat lng cay ieu qua cac nghien cu ve di truyen, chon giong sao cho cay phat trien nhanh, cho nhieu trai, hat to. Sau ay la mot vai so lieu ve san lng hat ieu qua mot so nam Bang 1: San lng hat ieu the gii qua mot so nam. n v : Tan. Nc Nam 1971 1975 1981 1989 1990 1991 1992 1993

Luan An Tot Nghiep

Braxin An o Mozabu Tanzania Girebitxao Kenia Indonexia Viet Nam Thai Lan Philipin Cac nc khac Toan gii the 15 177 175 175 175 23 23 23 23 23 23 505 60 166 185 185 185 30 30 30 30 30 5 586 75 185 70 70 70 15 15 15 15 15 5 410

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120 100 25 20 15 10 30 15 8 4 5 381 126 115 29 25 17 11 35 30 10 5 5 442 130 110 30 27 16 13 38 35 12 5 5 457 130 115 20 15 10 10 30 50 12 5 5 429 180 120 18 15 10 8 27 72.5 12 5 3 445. 2

Bang 2: Tnh hnh trong ieu STT 1 2 3 4 5 6 7 8 9 10 11 a phng An o Mozambie Braxin Tanzania Kenya Philipin Madagascar Nigieria Srilanca Malaysia Irorycoast Dien tch (ha) 150.000 150.000 80.000 58.000 50.000 4.200 2.000 1.000 800 600 600

(Nguon bao cao th trng mot so nong san cua Bo Thng Mai 1993). 7

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2. S lc ve nc trai ieu: oi tng khai thac che bien trai ieu la dch qua. Phan dch qua chiem tren 80% trong lng trai ieu. Theo nhng nghien cu trc ay tai Bo mon Cong nghe Thc Pham, trng ai hoc Bach Khoa Tp. Ho Ch Minh, thanh phan hoa hoc cua dch trai ieu c trnh bay bang sau:

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Bang 3: Thanh phan hoa hoc cua dch trai ieu mot so tnh mien Nam Hoc Mon a (TPHCM) phng Chat Chat (%) ng (%) kho 11.80 0.37 10.53 10.63 247 0.12 0.94 0.33 84.25 0.19 Long Khanh (ong Nai) 11.38 0.37 9.87 9.87 244 0.18 0.97 0.32 83.50 0.20 Thu Dau Mot (Song Be) 11.82 0.38 9.78 9.82 252 0.19 1.25 0.33 85.50 0.19 My Tho (Tien Giang) 11.85 0.39 10.40 10.05 268 0.20 1.01 0.22 84.10 0.19 Phan Thiet (Bnh Thuan) 11.92 0.36 40.48 10.05 249 0.18 1.01 0.33 83.40 0.19 Phan Rang (Ninh Thuan) 11.93 0.37 10.55 10.57 245 0.17 1.04 0.32 83.50 0.19 11.97 0.37 10.25 10.30 249 0.17 1.03 0.32 84.10 0.19 Trung bnh

Tannin (%) kh

ng tong (%) Vitamin C (mg %) am thanh phan (%) Protid (%) Tro (%) Nc (%) Acid citric (%) tho

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o pH 4.00

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4.10 4.00 4.10 4.10 4.10 4.03

10

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T bang so lieu tren, ta thay dch trai ieu cha thanh phan dinh dng ang quan tam la ng va vitamin C. Ngoai ra, tannin cung chiem ham lng kha cao; chnh thanh phan nay gay ra v chat ang cho nc ieu. Tannin con gay nhng tac ong bat li ve dinh dng, c che c the hap thu protein va gay tao bon. Thanh phan tannin la nguyen nhan chnh lam giam gia tr s dung cua trai ieu. Ham lng tannin trong nc trai ieu giam theo o chn cua trai ieu.

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ONG NAI CAC CH SO IEU VANG CX A. Nguyen trai 72.3 1:14 81.3 chat 8.5 0.56 8.5 8.5 238 76.5 1:14 81.0 10.5 0.35 9 9 241 0.17 4.56 0.23 80.0 1:16 81.8 11.5 0.32 9.45 9.45 244 0.18 4.25 0.22 60.0 1:14 82.5 9 0.55 8.9 8.9 239 0.15 4.44 0.28 65.0 1:14 80.5 11 0.36 9.4 9.4 239. 5 0.16 4.2 0.26 67.5 1:16 81.2 12.5 0.35 9.5 9.56 244 0.18 4.41 0.26 57.4 1:14 83.7 8.7 0.55 8.7 8.9 238 0.16 4.05 0.16 63.9 1:14 84 10.2 0.4 92 9.35 246 0.17 4.10 0.18 1. Trong lng trung bnh 2. T le hat/ trai B. Phan nac 3. o am (%) C. Dch ep trai ieu 4. 5. 6. 7. Ham lng kho (%) Ham (%) CT CR IEU O CX CT CR

SONG BE IEU VANG CX CT CR 65.3 1:16 85.5 11.8 0.38 9.78 9.82 252 0.19 4.00 0.19 IEU O CX 55.4 1:14 84.6 8.5 0.53 8.5 8.6 240 0.16 4.10 0.16 CT 61.9 1:14 84.9 10.3 0.39 9.0 9.15 248 0.17 4.15 0.17 CR 63.0 1:16 85.7 11.8 0.37 9.78 9.82 253 0.18 4.00 0.28

lng Tannin

Ham lng ng kh (%)

Ham lng ng 0.16 tong (%) 4.14 8. Ham lng vitamin 0.27 C (mg%) 9. Nit tong (%)

12

Luan An Tot Nghiep

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10. o pH 11. o acid (theo acid citric)

13

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3.

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Hien trang khai thac va che bien trai ieu: 3.1/ cac nc tren the gii:

Tren the gii, cac nc trong nhieu ieu ngay cang quan tam vao viec s dung trai ieu e san xuat cac loai nc uong khac nhau. Chang han nh nc giai khat trch t trai ieu co ten la Cashola. San pham nay co 290Bx va 4% acid. Nc ep t trai ieu c dung pha vi nc hoac sa va ng, c goi la Cajuda hoac san pham Cajuvita co cha nhieu vitamin Cajuasporative dung e pha vi mat ma. Goa (An o) ngoai cac san pham tren con co ru Arrack va Feni len men t nc trai ieu. Ngoai ra, trong trai co mot t pectin, rat thch hp e lam mt. Vao the ky 18, mt ieu kha pho bien Phap va t ay c lan sang cac nc Chau Au. Do kho khan trong viec thu gom, tnh khong ben cua trai ieu va viec tinh che dch qua ieu phc tap, nen du san lng trai ieu rat ln, nhng so lng a vao che bien khong nhieu. 3.2/ Tnh hnh che bien ieu tai Viet Nam: Trai ieu la mot loai san pham nong nghiep cha c khai thac co hieu qua tai Viet Nam. Theo thoi quen nhan dan ta thng dung trai ieu nh mot loai rau trong an uong nhng vi so lng khong nhieu. T 1980, khi co chu trng phat trien trong ieu e lay hat, ong thi cung at ra viec s dung che bien trai ieu, mot so c quan, n v nh: Vien Khoa hoc Lam nghiep Ha Noi, Trung tam Cong nghiep Sinh Hoc thuoc Phan vien Khoa hoc Viet Nam, Bo mon Cong nghe Thc pham trng ai hoc Bach Khoa Tp. Ho Ch Minh, co tien hanh nghien cu khai thac nc trai ieu va che bien thanh ru len men hoac ru ieu pha che... Tuy nhien pham vi s dung cua loai san pham nay kha hep, khong trien khai m rong c san xuat. Mong muon chung la phai a dang hoa san pham t nc trai ieu. V du nh nc uong, mt, banh keo... ma tr ngai ln nhat la nc trai ieu co ham lng tannin cao, gay v ang chat ma ky thuat loai bo tannin kha phc tap, cha tm c bien phap tach tannin n gian va co hieu qua.

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Quy mo s dung trai ieu vao che bien cong nghiep chu yeu la ky thuat loai bo tannin khoi nc trai ieu.

II. AC TNH CUA TANNIN THC VAT VA TANNIN TRONG NC TRAI IEU 1. ac tnh cua Tannin thc vat 1.1/ nh ngha, phan loai tannin. Phc chat tannin hay chat chat c hieu la mot hon hp phc tap cua nhieu hp chat co ac tnh la Polyphenol. Trong lng phan t t 500 en 3000 vC. Song qua nhng tai lieu c biet en. ngi ta a khang nh rang trong thanh phan cua tannin th flavan 3ol va flavan 3,4 diol chiem phan ln (khoang 85 92% tuy tng loai thc vat va tuy tng giong, loai) trong phc chat tannin.
OH HO O OH OH OH

Flavan 3ol

OH OH O OH OH OH OH

Flavan 3,4 diol

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Ngoai ra con co cac hp chat phenol n gian khac cung c biet en trong hp chat polyphenol nh:

OH OH HO

OH OH

Pirocatechin
OH OH HO

Pirogalol

O HO HO OH C OH

Florogluxin

a. Galie

O HO HO C O O OH O C OH

a.

ellagic

Cung vi nhieu phenol n gian khac. Theo cac tai lieu co c hien nay, ngi ta phan loai tannin theo 2 cach sau: Phan loai tannin thanh Pirocatesie va Pirogalie: Phng phap phan loai nay chu yeu da vao phan ng hoa hoc gia tannin va dung dch FeCl3. ong thi da vao kha nang phan giai tannin khi un nong no trong dung dch KOH thanh cac cau t khac nhau. - Neu la tannin pirocatesie th khi tac dung vi dung dch FeCl3 cho ket tua mau xanh nhat va khi un nong trong dung dch KOH se phan giai thanh Pirocatesol. 16

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- Neu la tannin Pirogalie khi tac dung vi dung dch FeCl3 cho ket tua mau xanh tham va khi un nong trong dung dch KOH se phan giai thanh pirogalol. Phan loai tannin thanh tannin thuy phan va tannin ngng tu. Phng phap phan loai nay da vao cau truc phan t, chu yeu da vao moi lien ket gia cac nhan benzen vi nhau trong phan t, ong thi da vao kha nang phan giai hay ngng tu cua tannin khi un nong trong dung dch acid hoac di tac dung cua enzym oxy hoa. Neu la tannin thuy phan: trong phan t cua no, cac nhan benzen lien ket vi nhau bang moi lien ket este phc tap, ong thi khi thuy phan se phan giai thanh cac cau t n gian. Cac cau t nay c phan giai ra t tannin thuy phan thng la cac loai ng monophenol va acid phenolcacboxylic. Chat tieu bieu cho tannin thuy phan c goi la tannin ac biet. Khi tac dung vi FeCl3 cho mau xanh nhat. No c tao thanh chu yeu do s ket hp gia cac loai ng vi galie hoac acid catosie theo moi lien ket este phc tap. Trong khi phan t cua acid galie co cha nhom phenol (-OH) va nhom cacboxyl (-COOH) cho nen gia 2 phan t acid galie co the ket hp vi nhau e tao thanh hp chat este phc tap goi la depxit. Neu 3 phan t ket hp vi nhau se tao thanh hp chat tridepxit v.v...

HO HO HO COOH +

OH HO OH HO COOH

HO COO HO

OH OH

COOH

Hai acid galie Hoac

acid metha - di-galie

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CH2 OK CH O [CHO K ]3 CHO K

Trong o K :
OH CO OH OCO OH OH OH

Neu la tannin ngng tu: cau tao phan t cua no khac han vi cau tao phan t cua tannin thuy phan. Trong phan t cua tannin ngng tu, cac nhan benzen lien ket vi nhau khong phai bang moi lien ket este phc tap ma bang moi lien ket cacbon trc tiep. Do o khi un nong trong dung dch acid loang, tannin ngng tu khong phan giai thanh cac cau t n gian, ngc lai se ngng tu thanh hp chat phan t ln. Khi tac dung vi Fe-acetate, tannin ngng tu cho chat mau xanh. Chat tieu bieu cho tannin ngng tu la Catechin va galocatechin

OH HO O OH OH HO OH HO O OH OH OH

Catechin

Galocatechin

Nhng nghien cu gan ay nhat ve cau tao cua catechin a xac nh c rang o la chat co cau tao cua flaranol-3 va hien nay cau tao cua 18

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flavanol-3 a c chon lam nen tang cua tat ca cac dang polyhydroxylflavan
2' 2 3 1' 3' 4' 5'

8 7 6 5

O
4

OH

6'

Flavan 3 - ol

HO O HO H OH OH HO OH OH

OH OH

OH

OH HO O OH OH

HO

OHOH

OH OH

OH

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S ngng tu cua catechin

OH HO OH OH

OH HO O OH OH OH
OH HO O

OH OH

OH OH O

OH

H+

OH HO NAD

(P)H
OH

O 4
3

OH OH

NAD (P)H HO O
OH H OH

OH OH

H+
OH

OH
OH

HO

O H OH

HO

O H OH

OH

OH

OH

OH HO O H OH OH

OH

C che d kien cua qua trnh sinh tong hp eatechin va procyanidine. 20

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1.2/ Cac tnh chat c ban cua tannin: Nh ta a biet, cac loai tannin lay t cac thc vat khac nhau. Tuy thanh phan to hp cua chung khong giong nhau nhng ngoai cac ac tnh rieng chung van co chung mot so tnh chat c ban. Tannin thc vat la mot hon hp phc tap cua cac hp chat phenol. Sau ay la mot so tnh chat c ban cua tannin thc vat noi chung. Trong lng phan t cua tannin bien thien t 500 3000. the ran, tannin la chat ket dnh hoac vo nh hnh co v chat ang mc o khac nhau. Trong y hoc, tannin cua mot so cay c dung lam thuoc. Trong cong nghiep, tannin c dung vao cong nghe thuoc da. Noi chung cac loai tannin eu tan trong nc (tr tannin ket hp, hoa tan c trong mot so dung moi hu c nh ru, ether ethyl, acetone, ethyl acetate. Phan ln tannin khong tan trong benzen, ether dau hoa va clo. Khi tac dung vi dung dch FeCl3, tannin tao thanh phc mau xanh nhat hay xanh tm phu thuoc vao so lng nhom (-OH) co trong phan t. Tannin tac dung vi dung dch kali bicromat hoac acid cromic tao thanh ket tua mau xanh nhat. Tannin tac dung vi vanilin tao thanh phc vaniline - tannin co mau nau o trong moi trng acid. Phc nay co kha nang hap thu cc ai bc song = 500 nm. Tac dung vi dung dch Amonferixianua, tannin tao thanh phc chat co mau o tham. Khi tac dung vi cac dung dch protein, alkaloit se tao thanh phc tng ng vi ket tua. Tnh chat nay cua tannin co y ngha quan trong cho chung c dung e ket tua protein trong cong nghe thuoc da, kh oc khi b nhiem oc alkaloit va khi can thiet co the ket tua loai bo tannin ra khoi dung dch. Tannin co tnh kh manh, trong khong kh no de dang b oxy hoa. ac biet trong moi trng kiem no b oxy hoa rat nhanh. San pham t oxy hoa cua tannin la nhng chat co mau o, sau o chuyen thanh mau sang en hoac nau tham. 21

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Tannin b oxy hoa sau sac va triet e khi tac dung vi KMnO4 trong moi trng acid hoac vi dung dch iod trong moi trng kiem. Li dung tnh chat nay e nh lng tannin co trong nguyen lieu thc vat. Tannin tao thanh ket tua mau vang tro vi dung dch acetate ch. Tnh chat nay dung e tach tannin ra khoi dch chiet thc vat hoac nham muc ch thu hoi tannin hoac loai bo tannin. Ngoai nhng tnh chat tren, tannin con co the tac dung vi nhieu hp chat vo c va hu c khac. ac biet di tac dung cua cac enzym oxy hoa kh tng ng, tannin thc vat de dang b oxy hoa va ngng tu thanh cac san pham co mau va khong mau, anh hng trc tiep en mau sac va cac ch tieu chat lng khac cua thc pham c che bien t nguyen lieu thc vat. 1.3/ Mot so phan ng thng gap cua 1.3.1/ Phan ng oxy hoa: Di tac dung cua enzym polyphenoloxydaza, cac polyphenol b oxy hoa e tao thanh cac quinon. Ban than cac octoquinon cung la nhng chat oxy hoa va neu co mat cac chat kh nh vitamin C, glucose, chung se oxy hoa cac chat o va tr lai thanh polyphenol theo s o sau.
Polyphenol 1/2O 2

tannin

AH2

Octoquinon

H2O

OH
8 2'

O OH HO
5' 8 2'

O B Y
6'

HO
6

O OR 6' OH

O A

OH

R 1 = X1 = H 22

R = X1

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R = X2

SV: Trng Tan Thong

R2 = X2 Y=H Y = OH
OH OH B Y O

2'

HO A
6

OH

O O OH

Y=H Y = OH

2'

OH OH

X2

C
6'

OH

S oxy hoa ket hp flanvanes - 3 - ol cua tra trong qua trnh hnh thanh Thearubigin trong he polyphenol oxy hoa. 1.3.2/ Phan ng trung hp oxy hoa cua

tannin:

O HO OH O HO O H OR OH OH

OH

OR2

OH OH OH OH OH

HO

OH

OH

OH HO O H OH R1 HO H OR2 O

OH

OR1

OH

23

OH

H hoa X c

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Cac loai tannin t tra en. Cac san pham trung hp dimer oxy hoa 1.3.3/ Phan ng vi cac protein khong tan, protein tan va protein bat ong:
T Psol T Psol Psol T Pin

+ T: tannin + Psol : Protein tan + Pin : Protein tan Cac moi quan he gia hp chat tannin va protein 1.3.4/ Phan ng cong: Khi co mat cac acid amin, cac octoquinon c tao thanh se tien hanh phan ng cong hp vi cac acid amin. Cac san pham c tao thanh cua phan ng cong hp lai co the b oxy hoa e tao thanh cac octoquinon tng ng. 1.3.5/ Phan ng ngng tu: Cac octoquinon de dang ngng tu vi nhau e tao nen cac san pham co mau gia v chung la flobaten Polyphenol + O2 polyphenoloxidaza octoquinon (1) Octoquinon + H2O (2) n (octoquinon) glucose Polyphenol + O2 + Flobaten (3)

Trong ieu kien ho hap bnh thng th phan ng (1) va (2) chiem u the. Trai lai trong ieu kien san xuat, khi cac mo va te bao thc vat b pha v th phan ng (1), (3) chiem u the, trong o phan ng (1) la qua trnh enzym va phan ng (2), (3) la qua trnh phi enzym. 24

Luan An Tot Nghiep

SV: Trng Tan Thong

S tao thanh phc tannin - protein phu thuoc chu yeu vao viec hnh thanh nhng lien ket hydrogen gia nhom chc phenol cua tannin va cac nhom chc lien ket peptid cua protein.
H N C O HO O OH NH O C CO NH OH CO

O H O C O H N H

Cac kieu trao oi lien ket hydrogen gia nhom phenol cua flavane 3-ol va chuoi polipeptid

( CH NH CO NH CH ) 2 2n
CH2 CH2

O
C O

CH2 N

H O C
n

O H N

CH CH2

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Luan An Tot Nghiep

SV: Trng Tan Thong

OH

H N C O HO R CH N H OH

NH OH O C CH R H OH N C O

Cac polimer tong hp ma no co the ket tua c tannin 2/ ac iem tannin trong nc trai ieu: Qua ket qua thc nghiem trong qua trnh nghien cu, toi co nhng ghi nhan bc au ve tannin trong nc trai ieu nh sau: Tannin trong nc trai ieu tan trong nc, ether, aceton, va ethylic. Tannin trong trai ieu cung tao phc mau tm xanh vi FeCl3. Tannin nay cung tao phc mau vang tro vi acetate ch. Khi tac dung vi dung dch Kali Bicromat cho ket tua mau xanh nhat. 4. Mot so phng phap tach tannin: (?) Trc ay a co nhieu nghien cu a ra viec tach tannin nh phng phap dung gelatin, bentonito, nha anion hay dung phng phap sieu loc... Trong luan van nay xin a ra hai phng phap tach tannin khac la: dung nha trao oi cation va dung silicagel. 4.1. Dung nha trao oi cation: Hau het cac hp chat flavonoid sau khi c acid hoa se chuyen sang dang muoi flavanium. Li dung ac tnh nay, ta cho dch chiet chay qua nha trao oi cation. Khi o cung vi cac ion dng khac, tannin (flavalium) se tien hanh phan ng trao oi vi nha va b gi lai tren cot nha cation. Sau o dung nc nguyen chat e ra cac hp chat hu c dang phan t va ng ra khoi cot. Tiep theo dung mot dung moi phan cc manh hn e thc hien phan ng trao oi vi nha cation (nha bay gi ion

26

Luan An Tot Nghiep

SV: Trng Tan Thong

di ong la tannin). Ket qua la dch tannin c thoat ra ngoai. 4.2/ Dung silicagel T lau ngi ta a s dung phng phap sac ky e lam tinh khiet cac hp chat thien nhien. ng dung phng phap sac ky cot e tach tannin ra khoi dch trch. Ta cho dch trch vi dung moi hoa tan la ether qua cot. Li dung s hap phu khac nhau cua cac cau t trong hon hp dch trch, sau o dung dung moi thch hp la ether ethyle e ra giai nham loai ng va mot so tap chat khac ra khoi dch trch qua cot. Tiep theo dung dung moi phan cc manh hn e tien hanh giai hap thu tannin. Ket qua nhan c tannin.

5. Cac yeu to anh hng en o ben cua tannin: So vi cac hp chat thien nhien, tannin la chat co o ben kem, no ch the hien tnh ben trong moi trng acid. Vi tac dung cua nhiet trong moi trng pH acid, no de b phan huy va sau o ngng tu lai voi nhau thanh cac hp chat co mau nau. Ngoai ra s phan huy cua tannin con co the xay ra trong qua trnh chiet, trch cung nh tinh che chung. o ben cua tannin phu thuoc vao nhieu yeu to nh cau truc hoa hoc cua tannin, pH, nhiet o, enzym. 5.1/ S chuyen hoa cac chat di tac dung cua pH va nhiet. Vao nam 1934, T.K. Seshachi, bang cac th nghiem vi cac tac nhan acid va nc nong, Ong a chng minh rang cac cacbon v tr so 2,4,8 lien ket vi hydro rat linh ong. ay la trung tam gay ra cac phan ng ngng tu cua catechin.

27

Luan An Tot Nghiep

SV: Trng Tan Thong

OH OH HO O OH HO O OH OH H+ H20
OH

HO

O OH OH

H+ H 2O

OH HO HO OH CH OH OH O OH OH OH OH OH

HO

O OH

OH
OH O OH OH OH

(C)

(D)
OH

Trong cong thc (B) va (C), lien ket mi c keo t nguyen to cacbon 2 cua mot catechin ti nguyen to cacbon 6 cua mot catechin khac. Nhng cung co lien ket khac c keo dai t v tr cacbon so 4 cua mot catechin ti v tr so 8 cua mot catechin... Tat ca cac lien ket o tao nen mot mang polymer e hnh thanh hp chat tannin co s ngng tu phc tap. Mayer va Merger a nhan thay rang trong nc nong hoac trong acid cac polyhydroxylflavan 3,4 diol trai qua hang loat s ngng tu.

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Luan An Tot Nghiep

SV: Trng Tan Thong

OH HO O OH OH OH OH OH OH OH

OH HO O OH OH OH OH OH OH O

-H2O

OH HO O OH OH OH OH OH OH HO + OH OH O OH HO O OH OH OH

Roux, qua nhng th nghiem a cho thay rang, di tac dung cua acid phlobaphene 3,4 hnh thanh mot lng ang ke arahocyanidene. Hau het cac phan ng hoa hoc lien quan en o ben tannin noi rieng va cac hp chat polyphenol noi chung, toc o thuy phan cua chung eu phu thuoc vao nhiet o. Tnh ben cua tannin phu thuoc vao cau truc cua chung, pH, s co mat cua oxy va s tac ong qua lai gia cac thanh phan.

R OH HO O
+

R' O HO K2 O ogl ogl R'

ogl ogl

29

Luan An Tot Nghiep

SV: Trng Tan Thong

HO

O ogl

R1 OH R2

ogl

gl : gluco Khi mot cau truc cua tannin ben vi s gia nhiet th ong thi cung ben vi s tang pH. S hydroxyl hoa cac agly lam giam hoat tnh ben cua tannin. Trong khi s metoxyl hoa aglycosyl, acyl hoa co o ben nhiet ln nhat tai pH = 1.8 . 5.2/ Tnh ben cua tannin phu thuoc vao s co mat cua oxy Oxy va nhiet o c xem la nhng tac nhan ac trng xuc tien s phan huy cua tannin, t o sinh ra nhng dang san pham khong mau hoac mau nau. Oxy se oxy hoa manh liet tannin khi nung nong. 5.3/ Tnh ben cua tannin phu thuoc vao

enzym:

Cac enzym oxy hoa kh- chu yeu la polyphenoloxidaza va peroxydaza co trong hau het dch ep trai cay. S chuyen hoa- di tac dung cua cac enzym peroxydaza co san trong ban than nguyen lieu tannin se b bien oi sau sac tao nen cac san pham oxy hoa tan va khong tan. Enzym polyphenoloxydaza gay ra s oxy hoa tannin co kem theo s tieu thu mot lng ln oxy lam giam ham lng chung cua hon hp tannin va tao ra cac san pham co mau. Di tac dung cua enzym peroxydaza, lng tannin giam i rat ln va gay ra hien tng ngng tu tannin mc o cao, ong thi lam giam ham lng cac cau t thap, chuyen tannin sang trang thai phan t ln tan va khong tan trong nc. S bien oi di tac dung cua peroxydaza khong tao ra cac chat mau. 30

Luan An Tot Nghiep

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III.

CAC TAC NHAN CHNH E LAM SACH TANNIN 1. Silicagel: Trong ky thuat, ngi ta chia silicagel ra lam 2 nhom: Nhom 1 : silicagel dung cho viec chong am. Nhom 2: silicagel dung lam chat hap phu xuc tac hay chat on polymer, ac biet dung cho viec phan tch sac ky. 1.1/ Silicagel dung chong am: Co thanh phan hoa hoc la SiO2.

c ieu che bang cach cho kali hoac Natrisilicate (thuy tinh long) cho tac dung acid trong dung dch nc. o gel cua poly acid silic c tao thanh. Sau o tach nc thu c silicagel co nhieu lo xop. Silicagel nay c dung trong ky thuat chong am. 1.2/ Silicagel dung cho sac ky: Cung co thanh phan hoa hoc SiO2, c ieu che bang cach thuy phan SiCl4 trong moi trng hi nc. S thuy phan trong ieu kien ac biet co the tao c khoi cha cac hat hnh cau kch thc nho, co khi nho en khoang 100A0 va sau o bo ngng tu, bot nhuyen SiO2 se tao thanh. Cac loai muoi trang nay c s dung rong rai lam chat hap phu, xuc tac, chat on cho vat lieu polymer va ac biet c ng dung cho viec phan tch bang phng phap sac ky. 2. Nha trao oi ion: Nha c cau tao bi hp chat cao phan t gom mot sn hydrocacbon, co mang nhng nhom chc hoat ong , noi vi ion linh ong bang lc hut tnh ien. Ion di ong tham gia can bang trao oi. Co hai loai nha chnh: 2.1/ Nha trao oi cation hay cationit: Nhom hoat ong la anion, ion di ong la cation tao nha co dang K-M+. Tuy dang hp chat cua nhom hoat ong, co nhng loai cationit sau:

31

Luan An Tot Nghiep

Nhom sulfonat:

SV: Trng Tan Thong

( CH2 CH ) n

SO3 H

- +

Nhom phosphat
CH2 CH

PO4 (M )2

Nhom carboxylat
CH2 CH

COO H

- +

Nhom amino diacetat

CH2 CH

CH2 N

CH2 COO M
-

+ +

CH2 COO M

2.2/ Nha trao oi anion hay anionit oi vi nha trao oi anion, nhom hoat ong la cation, ion di ong la anion, tao nha co dang K+X-. Nhom ammonium 32

Luan An Tot Nghiep

CH2 CH

SV: Trng Tan Thong

CH2 NH3 X

+ -

Ngoai ra, con loai nha vo c, hp chat co cau truc dang silico aluminat, co cau tao phc tap nhng cho ket qua kem, do o kem thong dung. Nha thng co dang hnh cau, ng knh khoang vai chuc m en 1mm, co nhieu ten thng mai khac nhau.

IV. MUC TIEU VA NOI DUNG NGHIEN CU: Muc tieu cua nghien cu trong luan van nay la a ra mot so phng phap trch, lam sach nhom polyphenol trong nc trai ieu e phuc vu cho viec xac nh cac thanh phan tao nen nhom polyphenol trong nc trai ieu thong qua cac noi dung sau. Bc au nghien cu kha nang trch tannin t ieu khi c say thang hoa vi cac dung moi hu c khac nhau. Tiep tuc thu nhan tannin t dch trch vi cac chat hap phu khac nhau va nha trao oi cation. Bc au xac nh cac thanh phan cua tannin bang sac ky giay.

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Luan An Tot Nghiep

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Phan 3:

Nguyen lieu va phng phap nghien cu

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I. NGUYEN LIEU: 1. Trai ieu:

SV: Trng Tan Thong

Nguyen lieu chnh s dung trong suot qua trnh nghien cu la trai ieu kho a c say thang hoa vi o am 22.6% va c bao quan lanh. 2. Cac chat dung e trch, lam sach tannin: 2.1/ Silicagel: Silicagel c s dung trong cac th nghiem la bot silicagel dung cho sac ky co c hat 15m, c san xuat Hungary. 2.2/ Nha trao oi cation: Trong luan van nay, nha trao oi cation c s dung la nha co nhom hoat ong la anion va ion di ong la cation - nha trao oi carboxylate.
CH2 CH

SO3 H

- +

Do cong ty Bayer cua c san xuat. 2.3/ Nhom oxyt Al2O3: Nhom oxyt dung trong cac th nghiem nay la Al2O3 c hat 15m do Phap san xuat. 3. Cac hoa chat: 3.1/ Cac dung moi: Dung cac dung moi: aceton, con 99.5%, ethyl ather, benzen, buthanol, acid acetic. 3.2/ Cac hoa chat khac: KMnO4 , HCl, Indigo Carmin, H2SO4 , FeCl3 , K3Fe(CN)6 , Pb(CH3COO-)2, NaOH, glucose.

II. PHNG PHAP NGHIEN CU 35

Luan An Tot Nghiep

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1. Chon dung moi trch tannin Bc au xac nh kha nang trch tannin cua cac dung moi sau trong moi trng acid pH=2. Ether ethylic Aceton Ethylic Nc : ieu kho : ieu kho : ieu kho : ieu kho 60ml : 30g 60ml : 30g 60ml : 30g 60ml : 30g

36

Luan An Tot Nghiep

SV: Trng Tan Thong

2. Tien hanh lam sach tannin: 2.1/ Tac nhan la silicagel Trai ieu S 30g O NGHIEN CU 60 ml C2H5OH : CH3COOH 85:15 V/V (CH3)2CO : CH3COOH 85:15 Ngam V/V H2O : CH3COOH 85:15 V/V (C2H5)2O : CH3COOH 85:15 Loc V/V 3 ngay em nhiet o phong

Ba ieu

Co ac

{ { o chan khong 60
t0 : 250C (C2H5)2O : 10ml

Hoa tan Hap phu Silicagel Dch ng va mot so tap chat khac Ra cot 1 (C2H5)2O Ra cot 2 C2H5OH Tannin

{ 30g Silicagel

cot cao : 20cm

Co ac Th nghiem 2.2/ Tac nhan la nha cation Sulfunate: S O NGHIEN CU

37

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Luan An Tot Nghiep

Ngam

Ba ieu

Loc

{ {

60 ml C H5OH : CH3COOH 85:15 SV:2Trng Tan Thong V/V (CH3)2CO : CH3COOH 85:25 V/V H2O : CH3COOH 85:25 V/V (C2H5)2O : CH3COOH 85:25 V/V 3 ngay em nhiet o phong

Co ac

o chan khong 60 t0 : 250C

Hoa tan Trai ieu Trao oi nha cation Nc, ng, acid Ba ieu Ngam Ra giai 1

H2O : 60ml

{ toc o : 0.5ml/ph
40g nha Nc : a.acetic 85:15 V/V 3 ngay em Nc catphong nhiet o 20% Izopropyl alcohol

Ra giai 2 Loc Co ac

Trao oi nha lan 1 = 1 V = V1

Trao oi nha Th lan 2 nghiem = 2 V = V2

Trao oi nha lan 8 = 8 V = V8

Trao oi nha lan 9 = 9 V = V9

3. Xac nh cac thong so trong qua trnh trao oi nha cation: ng, chat Ra giai NGHIEN CU S O hu c khac H2O lan 1 cung mot phan tannin khong kp trao Ra giai 2 Izopropyl alcohol oi (d tha) nh lng tannin 38 Xac nh dung lng Th trao oi

Luan An Tot Nghiep

SV: Trng Tan Thong

........

1 > 2 >...>8 > 9

Xac nh c , , thi gian t, V

4.

Xac nh cac thong so trong qua trnh hap thu Silicagel S O NGHIEN CU

Trai ieu

Ngam

{
39

Nc : a.acetic 85:15 V/V 3 ngay em nhiet o phong

ng, chat Ba hu c khac ieu

Loc

C2 2H 5)2O (CH5OH 2 Giai hapXac nh Tannin b nh lng 1

Luan An Tot Nghiep

SV: Trng Tan Thong

Hap phu 1 Hchieu cao = h1

Hap phu 2 Hchieu cao = h1

Hap phu 3 Hchieu cao = h1

Hap phu 4 Hchieu cao = h1

h1 > h2 > h4 > h3

Xac nh c H = h4

Xac nh c cac thong so

IV. PHAN TCH TANNIN BANG PHNG PHAP HOA HOC a) Hoa chat s dung: Dung dch KMnO4 0.1N Dung dch indigo carmine 0.5% trong 2.5%(V/V) acid sulfuric. Hoa tan 0.25g indigo carmin vi 25ml nc cat va them vao 1.5ml acid sulfuric am ac, sau o nh mc 100 ml va c dung dch ch th. b) Cach tien hanh: Lay 20 ml dch trch a lam trong, dung pipet hut lay 5ml indigo carmin 0.5%. Cho them 25ml nc cat. em chuan o bang dung dch KMnO4 0.1N cho en khi chuyen t mau xanh dng cua indigo carmin sang mau vang sang th dng. Ghi lai so ml dung dch KMnO4 0.1N a chuan o. Song song tien hanh nh lng vi 45 ml nc cat va 5ml indigo carmin. 40

Luan An Tot Nghiep

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Tnh t le tannin trong 20ml dch trch. C 1ml KMnO4 0.1N tng ng vi 4.157mg tannin. V. SAC KY GIAY: Cac yeu cau oi vi sac ky giay nh sau: tinh khiet hoa hoc, trung tnh ve phng dien hoa hoc hap phu, ong nhat ve t trong, bao am cho dung moi chuyen ong vi toc o nhat nh. Co nhieu phng phap sac ky giay: mot chieu, hai chieu, vong. ay toi s dung sac ky giay mot chieu i len do phng phap nay n gian nhat. Viec chon dung moi e trien khai tuy thuoc vao mau, co the chon he dung moi sau: BAW : n-Buthanol - acid acetic - Water BAW : n-Buthanol - acid acetic - Water BEW 10) : n-Buthanol -95% ethanol - Water (4 : 1 : 5) (6 : 1 : 2) (4 : 2 : 2.2) (30 : 3 :3 :

BBPW : n-Buthanol - Benzen - Pyridine - Water -HCl : Water - acid acetic - 12N HCl 1%HCl acid chlohydric - water TAW : Toluen - acid acetic - water MAW : Methanol - acid acetic - water

(82 : 15 : 3) (3 : 97) (4 : 1 : 5) (90 : 5 :5)

e ac trng cho s chuyen ong cua cac chat tren giay, ngi ta bieu th Rf nh sau: Rf = Toc o chuyen ong mot cau t Toc o chuyen ong tuyen pha ong

Trong luan van nay chung toi s dung he dung moi BAW lan lt vi cac t le sau: (4 : 1 : 5) ; (4 : 1 : 2.2) ; (14 : 1 : 5) ; (4 : 1 : 1) ; (6 : 1 : 2) Bnh tieu ban dung chay sac ky cao 30cm, ng knh 13cm. Giay c s dung la giay Whatman 1. Mau c cham cach mep di 1.5cm, lng dung moi vao cach vet cham la 0.6cm. 41

Luan An Tot Nghiep

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Hai vet cham cach nhau 2cm. S dung ong mao quan e cham mau. Mau a c co ac, cham nhieu lan tai vet. Vet loang cac mau khong qua 1 cm.

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VI. PHNG PHAP PHAN TCH BANG MAY SAC KY LONG CAO AP: 1. Gii thieu ve may sac ky long cao ap: Ke t khi ra i bat au vao nam 1970, may sac ky long cao ap (High Performance Liquid Chromatography, viet tac la HPLC) ngay cang tien en mot mc o cao ve s tinh vi. Hien nay sac ky long cao ap c s dung nh la mot he thong may moc hoan chnh, tr thanh mot phng phap thong thng trong hau het cac lnh vc phan tch hoa hoc va no tiep tuc phat trien khong ngng. Thiet b sac ky long cao ap chu yeu gom be cha dung moi, bm, au phun, cot, au do va he thong thu nhan tn hieu. Trong he thong HPLC, cot la con tim cua he thong, cot phai tr, co thanh phang, ong nhat tren be mat va co the chu c ap suat cao. HPLC dung e phan tch, tinh che va xac nh ham lng cua cac chat da tren sac ky o. oi vi moi nhom chat co mot che o chay may rieng nh la cot, deletor (au do), toc o dong, pha ong, pha tnh, bc song, ap suat. HPLC c ng dung kha rong rai trong y te, moi trng cong nghiep hoa hoc va thc pham. Rieng thc pham, thanh phan gom a so nhng chat kh bay hi nen phng phap HPLC co ng dung rong rai. Co the dung HPLC e phan tch phan ln cac thanh phan thc pham nh ng, acid amin, polyphenol, cac chat phu gia khac, cac loai vitamin, cac chat mau... Trong HPLC thanh phan cua dung moi hoac pha ong la mot trong nhng yeu to anh hng en s tach biet cac thanh phan trong dung dch a vao phan tch. Pha ong co the la cac dung moi hu c hoac nc. Viec nhan biet cac chat tren sac ky o chu yeu da vao cac gia tr lu (cac peak tren sac ky o) cua chung. Thi gian va dien tch lu la cac ai lng cua cac chat va ch phu thuoc vao he sac ky c la chon. Dung HPLC e phan tch nh lng th co 4 giai oan: - Tach sac ky (thu tn hieu deletor). - Chuyen tn hieu thanh so lieu (dien tch). - So sanh cac so lieu. - Giai thch cac so lieu. 43

Luan An Tot Nghiep

SV: Trng Tan Thong

Nhng thuan li va bat li cua HPLC:

a) Thuan li: - Tnh a dang cao ve viec chon pha chuyen ong. - a dang ve c che (gi lai, duy tr). - Khong pha huy au do. - Phun trc tiep phc hon hp. - Phan tch nhanh. - o chnh xac cao. - Co the noi vi cac loai cot khac nhau trong mot lan phan tch. b) Bat li: - Cac dung moi co tnh oc, o nht cao va co tnh tron lan. - Thieu mot au do a nang. - Thiet b co the b h hong do an mon. - Thiet b, dung moi at tien. 2. Phng phap phan tch bang HPLC: Bang HPLC ngi ta co the xac nh, phan tch mot chat co nong o vai ppm mot cach de dang neu co ay u chat chuan, cot, dung moi va chon ung che o van hanh may (van toc dong, ap suat, bc song...). Trong khi thc hien luan van nay, chung toi dung HPLC hieu LC-10AS do Shimadzu (Nhat) san xuat. May nay ch chay c mot he dung moi va ch co mot au bm, nen con rat nhieu han che trong viec phan tch mau. a) Hoa chat s dung: - Methanol - KH2PO4 - Cac chat chuan. b) Tien hanh: Che o van hanh may: = 1ml/phut = 280nm (polyphenol) Pmax = 300atm

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Luan An Tot Nghiep

Cot : 120 - 5C18

SV: Trng Tan Thong

Pha ong : Methanol : Nc = 25% : 75% Cac chat chuan: - Can : 0.5002g - 0.0010g chat chuan - nh mc len 20ml bang pha ong. - Bm vao may se thu c sac ky a chuan. Mau th nghiem: - Hut 0.2ml dch trai ieu cho vao bnh nh mc - nh mc len 20ml bang pha ong. - anh tan bot kh. - Loc - Bm vao may, se thu c sac ky o. X ly ket qua: Da vao thi gian lu va dien tch cua cac peak, so sanh. - Mau nc trai ieu va cac mau chuan. - Mau nc trai ieu trc va sau khi x ly tannin

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Luan An Tot Nghiep

SV: Trng Tan Thong

Phan 4:

KET QUA NGHIEN CU

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Luan An Tot Nghiep

SV: Trng Tan Thong

I. CAC THONG SO TOI U DUNG CHO QUA TRNH NGHIEN CU: 1. Thong so toi u cua qua trnh trao oi nha: Toc o chay toi u : 0.5ml/phut. Khoi lng nha trao oi : 40g. The tch dch trch qua cot : 62ml. Khoi lng tannin c giai ly : 66.5mg. Dch trch co ham lng tannin ban au 1,2 mg/ml. 2. Thong so toi u cua qua trnh hap phu Silicagel: Dct trch trc khi qua cot co ham lng tannin : 52mg/10ml Khoi lng Silicagel : 30g Chieu cao cot : 20cm. Toc o dong chay ra khoi cot : 04 - 05 giot/phut. Khoi lng tannin c giai hap: 49.5mg.

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SV: Trng Tan Thong

II. SO SANH CAC PHNG PHAP TRCH: Tat ca cac th nghiem eu c tien hanh vi: a) Dung moi chiet 30g ieu kho vi o am 21.06%. 60ml dung moi trch. 10ml CH3COOH 1%. Ngam trong 3 ngay nhiet o phong. Trao oi tren nha cation: Ham lng tannin trong dch chiet (mg) trung bnh trong 3 lan Nc Aceton Ether ethylic Ethylic Nhan xet: oi vi aceton va ethyl ethylic ..(?).... de dang dch trch. Rat thuan li cho qua trnh trao oi cation sau nay. Lng tannin thu nhan c 90.18% & 87.5%. Ru ethylic kho co ac hn so vi aceton va therethylic. Trong ieu kien phong th nghiem cua trng khong tien hanh c, nen qua trnh trao oi cation sau khong thuan li. Lng tannin thu nhan c ch 75%. oi vi nc ta khong can co ac v sau o neu co ac ta lai phai pha loang vi nc e thc hien qua trnh trao oi nha cation. Lng tannin thu nhan la 94.23%. Co the mot so nhom polyphenol tan trong nc, aceton, eher ethylic, ethylic a b nha hap thu ma dung moi ra giai khong ra het c. 52 56 64 72 Ham lng tannin trong dch sau khi ra giai (mg) trung bnh trong 3 lan 49.0 50.5 59.0 54.0

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b) Hap thu tren cot Silicagel: Trong qua trnh hap phu tren silicagel, dch trch bang nc khong thc hien c do khong the co ac dch trch trong ieu kien th nghiem cua phong th nghiem hien co. Dung moi trch Ham lng tannin trong dch chiet (mg) trung bnh trong 3 lan Aceton Ether ethylic Ethylic 56 64 72 Ham lng tannin trong dch sau khi ra giai (mg) trung bnh trong 3 lan 54.5 63 69.5

Nhan xet: Ham lng tannin thu nhan c oi vi: Aceton Ethylic : 97.32%. : 96.52%. Ether ethylic : 98.44%.

II. SAC KY GIAY: Trong luan van nay, he dung moi trong khai trien sac ky giay la BAW, thi gian trien khai 8gi 30phut. Trien khai mau chuan ta ghi nhan c nh sau Dung moi dung cho pha ong: + n Buthanol - a. Acetic - Water (1) + n Buthanol - a. Acetic - Water (2) + n Buthanol - a. Acetic - Water (3) + n Buthanol - a. Acetic - Water (4) + n Buthanol - a. Acetic - Water (5) = (4 : 1 : 5) = (4 : 1 : 2.2) = (14 : 1 : 5) = (4 : 1 : 1) = (6 : 1 : 2)

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Luan An Tot Nghiep

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Hp chat phenolic chuan Catechin Protocatechi n Caffeic Hydrobenzoic P. Coumaric Ferulic Galie Tranciranavie Chlorogenic

Rf - 3

Rf - 2

Rf - 1

Rf - 4

Rf - 5

0.68 0.90 0.94 0.94 0.62 0.87 0.64 0.79 0.9

0.56 0.77 0.62 0.91 0.84 0.72 0.62 0.76 0.62

0.78 0.82 0.92 1 0.81 0.93 1 0.62 0.81

0.72 0.92 0.92 0.97 0.97 0.89 0.61 0.83 0.64

0.47 0.79 0.85 0.86 0.85 0.77 0.59 0.69 0.56

Nhan xet: Vi he dung moi so 3 (14 : 1 : 5): hon hp phenolic co Rf khac nhau kha xa so vi cac he dung moi khac. Chng to he dung moi nay lam cho hon hp phenolic phan tach tot hn cac dung moi (1), (2), (4), (5). Ket qua : Dung moi trch chon - ether ethylic. ..... dung cho sac ky giay : BAW (14:1:5).

b) Sac ky giay cho mau (14 : 1 : 5) Trong luan van nay, he dung moi trong khai trien sac ky giay chu yeu la BAW, thi gian khai trien: 8gi 30 phut. sau: Khai trien mau tren sac ky giay ta ghi nhan c nh

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+ Ch co hai vet tren giay sac ky, ngoai ra khong co vet nao khac. ieu nay cho nhan xet rang : trong dch trch cha hai cau t polyphenol hoac la co cha hai nhom cau t polyphenol ma trong moi nhom lai co cha cac cau t khac co Rf gan giong nhau va giong vi cau t chnh, va cung co khi la lng rat be so vi cau t chnh nen ta khong nhan c tren giay sac ky.

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Hnh .... : Sac ky giay

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Ta tnh Rf e xac nh cau t chnh. - Chieu cao di chuyen cua mau: + Vet gan nhat + Vet xa nhat : 1 - Chieu cao di chuyen cua dung moi: 21 Rf1 = 11/21 = 0.52 Rf2 = 14/21 = 0.66 Vi gia tr Rf tren, ta co the ket luan s bo trong dch trch co cac polyphenol sau: + P. Coumaric + Galie + Catechin : 0.62 : 0.64 : 0.68 :1

Va cung co the cung mot so phenolic khac nhng ay chung toi khong co mau chuan e so sanh. e co the ket luan chnh xac, ta co the tien hanh chay mau tren sac ky long cao ap. c) Sac ky long cao ap:

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