TRNG I HC DUY TNKHOA DC

Mn hc : Thc hnh ha sinh cn bn

Nhm : 2Lp : BCH201F1Thnh vin : 1/ Nguyn Hu an2/ Nguyn Th Thu Loan3/ on Th Ngc Ly4/ L Th Mi5/ Dng Th Mai

Nm 2014 2015MC LCBI 1: KHO ST ENZYMEBI 2: XC NH HOT CA ENZYMEBI 3: SACCHARIDEBI 4: LIPIDBI 5: AMINO ACID V PROTEINBI 6: TM NHNG CHT BT THNG TRONG NC TIU

BI 1:KHO ST ENZYMEI/ MC TIU:1. Nm c c ch hot ng ca enzyme.2. Bit c mt s enzyme thy phn c bn v nhng c cht tng ng c thy phn bi cc enzyme .

II/ THC HNH:1. Th nghim 1: Thy phn tinh bt bng Amylasea. Nguyn tc: Tinh bt khi c mt ca Amylase b thy phn thnh cc dextrin. Thi gian thy phn cng ko di th cng to cc dextrin vi phn t lng cng nh v cui cng l maltose. Cc sn phm to thnh mc thy phn khc nhau t tinh bt to thnh mu khc nhau vi dung dch It.b. Ho cht v dng c: ng nghim -Nc ct Cc c m-Nc bt ng ht-H tinh btc. Tin hnh:Lc u1 ml nc btDung dch nc bt 1/2019 ml nc ct

Cc c m

1 ml dd h 4tinh bt 1% + 0.5 ml dd nc bt 1/20 (ln lt mi ng)

2ng 2: 5p51ng 5: 20png 4: 15png 3: 10png 1: 0pSau thi gianNh 1 git dd It 1%34

d. Kt qu v bn lun: Quan st ta thy: ( t tri sang phi) ng 1: c mu tm than ng 2: c mu tm than nhng nht hn ng 1 ng 3: c mu nht hn ng 2 (mu nu) ng 4: c mu vng cam ng 5: c mu vng nht

Gii thch: Ti thi gian 0 pht, nh mt git It 1% vo ng 1, ta quan st c ng 1 c mu tm than. V 5 pht sau nh tip mt git It 1% vo ng 2, lc ny ng 2 s c mu nht hn ng 1. Tip tc nh vy 5 pht sau, li nh mt git It 1% vo ng 3, ng s c mu nht hn ng 2 (dn chuyn qua mu nu). ba ng 1, 2, 3: thi gian t 0 n 10 pht mu cn ti l v lng tinh bt cha c thy phn ht, cn nhiu nn khi cho It tc dng vi tinh bt cho ra mu nht dn t tm n nu. Vn nh vy, sau 5 pht, khi sang ng 4, ng s c mu vng cam, s chuyn i mu r rt hn v lng tinh bt b thy phn gn ht. Cui cng l ng 5, lng tinh bt b thy phn ht nn c mu vng nht.

2. Th nghim 2:Thyphn Ur bng Ureasea. Nguyn tc:Urease

CO(NH2)2+ H2O CO2 + 2NH3NH3 lm kim ha mi trng, vi s c mt ca phenolphthalein lm dung dch chuyn mu hng.b. Ha cht v dng c: ng nghim -Dd Ur 10% ng ht-Dd phenolphthalein 1%-Bt u nnhc. Tin hnh:Cho 2 ng nghim:1.5 ml dd Ur 10%

120.5g bt u nnh 1.5 ml dd phenolphthalein 1% 1.5 ml dd Ur 10% 1.5 ml dd phenolphthalein 1%

Lc u (5 10 pht)

d. Kt qu v bn lun: Quan st ta thy (t tri sang phi): ng 1: c mu pht hng ng 2: tch lm 2 lp: lp trn c mu hng b vn c, lp di c mu vng

Gii thch: ng 1, khi cho 1.5 ml dd Ur 10% v 2 git dd Phenolphthalein 1% tc dng vi nhau th ta thy xut hin m pht hng l bi v Ur c tnh kim yu nn khi cho phenolphthalein vo s c mu nh vy. ng 2, vn vi lng dd Ur 10% v dd Phenolphthalein 1% nh ng 1, v thm 0.5g bt u nnh th ta thy s tch lp, v Ur b thy phn bi bt u nnh to ra NH3(mi trng kim). Khi cho phenolphthalein vo th dd s c mu hng b vn c lp trn, lp di s c mu vng l do bt u nnh cha tan ht.

3. Th nghim 3:Thy phn Lipid bng Lipasea. Nguyn tc: Lipid (Triglyceride) b thy phn thnh glycerol v acid bo, s gii phng ra acid lm acid ha mi trng phn ng.b. Ha cht v dng c:

ng nghim -Dd sa nh tng ng ht-Dd phenolphthalein 1% -Dd Na2CO3-Nc ct -Dch tyc. Tin hnh:Cho 2 ng nghim:23 git dch tyNh t t dd Na2CO3 1%2 git dd phenolphthalein 1% 1 ml dd sa nh tng 3 git nc ctNh t t dd Na2CO3 1%2 git dd phenolphthalein 1% 1 ml dd sa nh tng 1

30 pht

d. Kt qu v bn lun: Quan st ta thy (t tri sang phi): ng 1: c mu hng ng 2: c mu trng c

Gii thch: Trong sa nh tng c lipid nn khi cho phenolphthalein vado th c hai ng u khng i mu. n khi nh t t Na2CO3 vo hai ng dd bt mu hng (v Na2CO3c mi trng kim). Tip theo khi cho nc ct vo ng 1 v dch ty vo ng 2, th dd trong ng 1 vn khng i mu, cn ng 2: v trong dch ty c enzyme thy phn lipid thnh glycerol v acid bo (mi trng acid), acid s tc dng vi base to ra mui (mi trng trung tnh) nn s lm mt mu hng

HNH NH C BA TH NGHIM

BI 2:XC INH HAT ENZYMEI/. MC TIU:1. Nm c phng php o quan v ng dng.2. nh lng sGOT, sGPT, amylase.

II/ THC HNH:CCH VN HNH MY SH PRIETEST TOUCH bi th nghim ny th c 3 th nghim u c nhng bc tin hnh ging nhau, ch khc t l pha mu th: i vi th nghim xc nh hot sGOT, sGPT: + Thuc th : 500 l+ Huyt thanh: 50 l i vi th nghim xc nh amylase huyt thanh:+ Thuc th: 500 l+ Huyt thanh: 5 lBc 1: Khi ng my V sinh my, kim tra dy cp in, ng dn ha cht sn sng. Bt cng tt ngun. H thng khi ng xong mn hnh chnh hin th MENU. Ra my: a nc ct n v tr ht mu, chn WASH ( tng t lp li 3 ln)Bc 2: Chy mu th bnh nhn Chun b mu bnh nhn (ht ha cht, huyt tng, ) Trn mn hnh chnh MENU di chuyn con tr n xt nghim cn lm (sGOT, sGPT, Amylase) sau chn ENTER chn SEL chn RUN Ch mn hnh hin th Press ASP Switch to sip a nc ct n v tr ht mu, nhn nt bn di ng ht.Sau ty loi xt nghim mn hnh s hin th mt trong hai trng hp sau:TH1: Mn hnh hin th Use stored result?, chn Yes. Ch mn hnh hin th Use blank stored result? chn Yes. Ch mn hnh hin th Read sample, a mu bnh nhn n v tr ht mu, nhn nt bn di ng ht. Kt qu s hin ra trong 3 pht.TH2: Mn hnh hin th Read sample, a mu bnh nhn n v tr ht mu, nhn nt bn di ng ht. Kt qu s hin ra trong 3 pht.Ch : + Sau mi loi xt nghim ra nc ct li trc khi lm xt nghim khc.+ Bm ESC tr v mn hnh chnh MENU.+ V sinh my sau mi ln th nghim.

1. Th nghim 1:Xc nh hot sGOT (ASAT) v sGPT (ALAT)a. Nguyn tc: ASAT (GOT) c xc nh da trn phn ng:L-Aspartate + -Ketoglutarate Oxaloacetate + L-GlutamateOxaloacetate + NADHH+L-Malate + NAD+ MDH Malate dehydrogenase ALAT (GPT) c xc nh da trn phn ng : Alanine + -Ketoglutarate Pyruvate + L-GlutamatePyruvate + NADHH L-Latate + NAD LDH Latate dehydrogenaseb. Dng c v ha cht: Ha cht. Huyt tng c li tm thnh huyt thanh. Thuc th Sgot v sgop. Dng c. ng nghim thy tinh. ng ht thuc th v huyt thanh. My o quang.

c. Tin hnh: nh lng hot sGOT trong huyt thanh. o bc sng 340nm (334 365nm) Nhit : 30 37C. c i chiu vi nc ct. Trong ng nghim cho vo:+ Thuc th : 500 l.+ Huyt thanh : 50 l. Trn u v sau khi 1 pht, c gi tr. nh lng hot sGPT trong huyt thanh. o bc sng 340nm (334 365nm). Nhit : 30 37C. c i chiu vi nc ct. Trong ng nghim cho vo:+ Thuc th : 500l.+ Huyt thanh : 50l. Trn u v sau khi 1 pht, c gi tr.d. Kt qu v bn lun: nh lng hot sGOT trong huyt thanh:* Kt qu:- V cAnh : 30,48 U/L.

- Kiu My : 22,43 U/L.

*Bin lun:- i vi V c Anh, tr s 30,48 U/L l bnh thng v tr s sGOT bnh thng i vi nam l t 13-46 U/L. Nu sGOT nh hn 13 U/L hoc ln hn 46 U/L th tr s sGOT l khng bnh thng.- i vi Kiu My, tr s 22,43 U/L l bnh thng v tr s sGOT bnh thng i vi n l t 11-39 U/L. Nu sGOT nh hn 11U/L hoc ln hn 39 U/L th tr s sGOT l khng bnh thng. nh lng hot sGPT trong huyt thanh:*Kt qu:- V c Anh : 24,21 U/L.

-Kiu My : 15,44 U/L.

*Bin lun:- i vi V c Anh, tr s 24,21 U/L l bnh thng v tr s sGPT bnh thng i vi nam l t 11-59 U/L. Nu tr s sGPT nh hn 11 U/L hoc ln hn 59 U/L l khng bnh thng.- i vi Kiu My, tr s 15,44 U/L l bnh thng v tr s sGPT bnh thng i vi n l t 9-37 U/L. Nu tr s sGPT nh hn 9 U/L hoc ln hn 37 U/L l khng bnh thng.T kt qu tn ta c c t s Deritis (ASAT/ALAT).- V c Anh : - Kiu My : e. Nhn nh kt qu:- Hot ezyme sGOT v sGPT trong huyt thanh ca V c Anh v Kiu My l bnh thng. - Trong mt s trng hp sgop v sgtp tng qu cao, qu thp hoc 2 tr s chnh lch nhau qu nhiu th c th mc 1 s bnh nh : vim gan cp tnh, vim gan mn, x gan, nhi mu c tim, tng trng lng dng c xng, vim xng, tiu myglobin.* Nhng yu t nh hng n tr s sGOT v sGPT trong huyt thanh.- Yu t gy gim: thiu vitamin B6.- Yu t gy tng: thuc iu tr tiu ng, gim au, chng thng phong,tr tim, chng ng kinh, nga thai.

2. Th nghim 2:Xc nh amylase huyt thanha. Nguyn tc: amylase

5CNPG3 3CNP + 2CNPG2 + 3G3 +2GCNPG3 : 2-chloro-4-nitrophenyl-maltotriosideCNP: 2-chloro-4-nitrophenolb. Dng c v ha cht: Ha cht. Huyt tng c li tm thnh huyt thanh. Enzyme amylase Dng c. ng nghim thy tinh. ng ht thuc th v huyt thanh. My o quang.c. Tin hnh: Thuc th: dung dch RGT pha sn Mu th: huyt thanh Bc sng o:400 410 nm Nhit : 25C - 37C. c i chiu vi nc ct. Trong ng nghim cho vo:+ Thuc th : 500 l.+ Huyt thanh : 5 l. Trn u, cho vo my o v c gi tr sau 3p.d. Kt qu v bn lun:*Kt qu:- V c Anh: 272,5 U/L

Kiu My: 274,8 U/L

*Bin lun:- i vi V c Anh, gi tr amylase huyt thanh cao hn so vi gi tr bnh thng ( >220 U/L).- i vi Kiu My, gi tr amylase huyt thanh cng cao hn so vi gi tr bnh thng ( >220 U/L).e. Nhn nh kt qu: Gi tr bnh thng: Amylase huyt thanh, huyt tng fructose > lactose > saccarose > formol nn khi h Cu2+c trong Fehling thnh Cu+mu m dn t ng 1 n 5.Sau khi un nng CuOH to thnh Cu2O c mu gchR-CHO + 2 Cu(OH)2R-COOH + 2CuOH + 2H2O2CuOH Cu2O + H2O2. Th nghim 2: Phn ng Furfural (Phn ng Molisch)a. Nguyn tc:-Nhng nhom OH ru bc 2 trong phn t ng co th bi kh co th kh nc bi axit manh nh HCl, H2SO4 m c tao ni i. -Nhng cht thu c nay la dn xut cua furfural, vi s co mt cua polyphenol tao thanh phc hp co mau c trng khac nhau.b. Dng c v ha cht:- Thuc th Molisch-ng nghim-Glucose 5%-ng ht-Fructose 5%-Lactose 5%-Saccarose 5%-H2SO4 m cc. Tin hnh:-Cho vo ng nghim 1ml mt dung dich saccharide, thm vao 2 giot thuc th Molisch. -Sau o nghing thanh ng, cho t t theo doc thanh ng nghim khoang 1ml H2SO4 m c. -Phan ng dng tinh khi xut hin vong tron mau tim mt phn cach gia 2 lp cht long.d. Kt qu v bn lun:-ng Saccarose: chia thnh 2 lp, gia c 1 vng mu xanh tm b mt phn cch gia hai lp cht lng, lp trn khng mu, lp di c mu tm.

Trc khi cho axit:

Sau khi cho axit:

-ng Glucose: chia thnh 2 lp, gia c 1 vng mu tm nht phn cch gia 2 lp cht lng, lp di mu pht tm, lp trn khng mu.

Trc khi cho axit:

Sau khi cho axit:

-ng Fructose: chia thnh 2 lp, lp trn khng mu, lp di xanh tm.Trc khi cho axit:

Sau khi cho axit:-ng Lactose: gia c 1 vng mu tm b mt phn cch 2 lp cht lng, lp trn muTrc khi cho axit:

Sau khi cho axit:

Gii thch:- Khi cho H2SO4 thi cac nhom OH ru bc 2 cua glucose, fructose, lactose, saccarose, formol se bi kh nc tao thanh furfural hoc chuyn hoa cht furfural.-Furfural phan ng vi cac polyphenol (Molish) cho ra nhung mau c trng theo tng loai ng.3. Th nghim 3: Kho st Polysacccharide (Kho st tinh bt)a. Nguyn tc:-Tinh bt khng tan trong nc lnh, trong nc nng to thnh dung dch keo gi l h tinh bt.-H tinh bt cho phn ng Molisch, khppng cho phn ng kh, vi It to dd mu xanh tm.b. Dng c v ha cht:- H tinh bt-ng nghim-Dung dch It-ng ht-My cch thyc. Tin hnh:Tinh bt vi It: Cho vo ng nghim 1 ml h tinh bt, thm vi git It, quan st mu xut hin, un si vi pht (un cch thy), quan st hin tng, sau lm lnh di vi nc, quan st.d. Kt qu v bn lun:Khi cho It vo tinh bt c mu tm than, sau khi un si dung dch mt mu, dung dch keo lm lnh di vi nc th dd li c mu tm than, nhng nht hn lc ban u.Ban u:

Sau khi un cch thy:

Lm lnh di vi nc:

Gii thch:-Trong tinh bt c amylose v amylosepectin, amylose s bt mu xanh vi It, amylosepectin bt mu tm vi It nn dung dch c mu tm than.-Mt khc, khi un nng th amylose b mt mu v tan trong nc nng, to keo v amylosepectin khng tan trong nc nng, v khi ngui th amylose s hin mu tr li.4. Th nghim 4: nh lng glucose bng phng php dng enzyme (Glucose Oxydase)a. Nguyn tc:-Glucose Oxidase (GOD) oxy ha glucose thnh gluconic acid v peroxide hydrogen (H2O2).-Peroxide hydrogen to thnh b enzyme peroxidase (POD) phn hy v gii phng oxy.-Oxy gii phng oxy ha 4-aminophenazone v phenol to phc cht quinonimine c mu hng.-Cng mu t l vi hm lng glucose. b. Dng c v ha cht: Ha cht. Huyt tng c li tm thnh huyt thanh. Thuc th Dng c. ng nghim thy tinh. ng ht thuc th v huyt thanh. My o quang.c. Tin hnh:Trong ng nghim cho vo:+ Thuc th : 500 l.+ Huyt thanh : 5 l.Trn u, 37C trong 5 pht.d. Kt qu v bn lun:Glucose trong huyt thanh ca Mai l 57.53 mg/dl

Gi tr glucose trong huyt thanh ca mai b hn gi tr bnh thng ( 4.1 6.4 mmol/l)

e. Nhn nh kt qu:- C th do nhn i lu ngy- Gim trong thiu nng tuyn yn, tuyn thng thn, dng nhiu insulin.

BI 4 + 5:LIPID, AMINO ACID V PROTEINI/ MC TIU:1. Kho st tnh ha tan ca Lipid trong cc dung mi khc nhau2. Nm c tnh cht ha hc ca amino acid, protein3. Hiu r v tin hnh c phng php nh lng protein, creatinine

II. THC HNH:1. Th nghim 1:nh lng triglyceride huyt thanha. Nguyn tc: Thy phn Triglyceride bng enzyme, xc ingj glycerol c gii phng bng phng php so mu:Lipoprotein lipase

Triglyceride + 3H2O Glycerol + 3R-COOHGlycerol kynase

Glycerol + ATP Glycerol-3-Phosphate + ADPGlycerol-3-Phosphate Oxydase

Glycerol-3-Phosphate Dihydroxyacetone Phosphate + H2O2PeroxydaseOxydase

2H2O2 + 4-amoniantipyrin + phenol quinone + 4H2O Thnh phn cc lipoprotein mu:LipoproteinPhn on LipidPhn onprotein

CholesterolTriglyceridPhospholipid

apolipoprotein

Chylomicron5%90%3%2%

VLDL15%60%15%10%

LDL45%10%20%25% (98% Apo B)

HDL17%8%25%50%(67% Apo A1, 22% Apo A2)

Cc th vi cht (Chymomocrom) VLDL: Lipoprotein c t trng rt thp [Very Low Density Lipoprotein] LDL: Lipoprotein c t trng thp [Low Density Lipoprotein] HDL: Lipoprotein c t trng cao [High Density Lipoprotein]b. Tin hnh: Trong ng nghim cho vo: Thuc th: 500 l Huyt thanh: 5 l Trn u v c gi tr sau khi 5 phtc. Kt qu v bn lun:Lng triglycerid: 132.1 (mg/DL) x 0.0114 = 1.50594 (mmol/l)

Lng Trigyceride cao hn so vi lng Triglycerid bnh thng

d. Nhn nh kt qu:

Loi m trong muTr s bnh thngTr s khng tt gy hi cho sc khe

Cholesterol ton phn6,2 mmol/L)

LDL Cholesterol4,1 mmol/L)

Triglyceride50 mg/dL>1,3 mmol/L)