Gc > Mc:Cao ng - i hc > Mc:Sinh hc > a57923a > a57928a > title:Thc hnh Sinh ha date:25-11-2010 sender:Trn Th Tho

Trang source:Trn Th Tho Trang type:ppt A. PHN NG MU CA PROTEIN I.1. Cch lm: ng nghim 1: cho vi 1 t tinh th biure, un nng chy cho n khi bt u cng li th ngng un. ngui, thm vo 1ml dd NaOH 10%, lc tan, cho thm vi git CuSO4 1%. - ng nghim 2: cho vo 2ml dd protein trng, thm vo 1ml dd NaOH 10%, lc tan, cho thm vi git CuSO4 1%. I. PHN NG BIUR I.2. Quan st: ng nghim 1: dd c mu tm hng - ng nghim 2: dd c mu tm xanh I.3. Gii thch: ng nghim 1: Khi un, 2 phn t ure kt hp li to thnh biure, lm xut hin lin kt peptit: -CO-NH-. Trong mi trng kim, biure kt hp vi CuSO4 cho phc hp mui Cu c mu tm hng. S to thnh biure: -----------------> t0 H2N -CO-NH2 + H2N -CO-NH2 Biure ure + NH3 H2N -CO-NH-CO-NH2 <--------> H2N -CO-NH-CO-NH2 HO -C-NH-CO-NH2 H bin = NH Mui phc hp ca Cu vi biure: + NaOH + Cu(OH)2 Phc hp mui Cu vi biure c m tm hng + Na+ + 4H2O

HN Cu 2(HO -C-NH-CO-NH2) = NH NH CO- =NH CO -NH NH-CO NH=COng nghim 2: Trong dd protein trng c lin kt peptit: -CO-NH- nn cng cho phn ng biure to phc hp mui Cu vi polypeptit c mu tm xanh. Mui phc hp ca Cu vi biure: + NaOH + Cu(OH)2 Phc hp mui Cu vi biure c m tm xanh + Na+ + 4H2O 2(HN-CH -CO-NH-CO-NH-CH-CO) R1 HN-CH-CO- =N-CH-CO-N-CH-CO R2 R1 R2 R3 HN-CH-CO =N-CH-CO- =N-CH-CO Ra Rb Rc Cu I.4. Kt lun: Phn ng biure l phn ng mu c trng pht hin lin kt peptit. tm ca phn ng khc nhau ty theo di ca mch polypeptit v lng muoiis CuSO4 II.1. Cch lm: Cho vo ng nghim 1ml dd protein trng, thm vo 1ml dd ninhydrin 0,1%, un si 1-2 pht. ngui.

II.2. Quan st: Dung dch khi un nng v lc ngui c mu tm m. II. PHN NG NINHYDRIN II.3. Gii thch Cc axit amin v cc peptit khi phn ng vi ninhydrin s b dezamin ha, oxy ha v decacboxy ha to NH3, CO2, v aldehyt tng ng. =O + =O =O H2N-CH -COOH R1 + H2O -OH -OH =O + NH3 + CO2 + R-CHO Ninhydrin dng oxi ha Ninhydrin dng kh =O + =O =O -OH -OH =O + NH3 Ninhydrin dng oxi ha Ninhydrin dng kh -OH -N= =O O= =O Sn phm ngng kt mu tm II.4. Kt lun: Phn ng ninhydrin l phn ng c trng pht hin nhm -amin. III.1. Cch lm: ng nghim 1: cho 2ml dd gelatin 1%, 1ml HNO3 un cn thn cho n khi bt trn ln thnh ng nghim. ngui, rt theo thnh ng nghim 1ml dd NaOH 10%. - ng nghim 2: cho vo 2ml dd protein trng, 1ml HNO3 un ln thy xut hin kt ta

v khi kt ta trn ln thnh ng nghim th dng li. ngui, rt theo thnh ng nghim 1ml dd NaOH 10%. III. PHN NG XANTO PROTEIN III.2. Quan st: ng nghim 1: khng i mu chng t phn ng khng xy ra. - ng nghim 2: khi un dd c mu vng ca dn xut nitro. Khi thm kim, sn phm ny chuyn thnh mui c mu vng da cam c trng. III.3. Gii thch - ng nghim 1: gelatin khng c axit amin nhn thm nn phn ng khng xy ra. - ng nghim 2: Khi cho HNO3 vo protein trng, cc axit amin nhn thm c trong protein trng s b nitro ha nhn thm to dn xut nitro c mu vng. NO2 CH2-CH -COOH NH2 Tirozin t0 + 2HNO3 OH CH2-CH -COOH NH2 OH O2N Dinitrotirozin (mu vng) H bin CH2-CH -COOH O2N NH2 =O =N =O - OH Th quinoit ca dinitrotirozin - Sn phm nitro ha tc dng vi kim to mui c mu vng da cam c trng. CH2-CH -COOH

O2N NH2 =O =N =O - OH CH2-CH -COOH O2N NH2 =O =N =O - ONa + NaOH + H2O Th quinoit ca dinitrotirozin Mui Na ca dinitrotirozin c cu to quinoit 9 vng da cam) III.4. Kt lun: Phn ng Xanto protein l phn ng c trng pht hin axit amin nhn thm. IV.1. Cch lm: ng nghim 1: cho 1ml dd protein trng, 1ml axit axetic h nng nh thy kt ta, sau un cho tan ht kt ta, lm lnh. Cho t t theo thnh ng nghim 2ml H2SO4 - ng nghim 2: cho1ml dd protein trng, 2 git saccaroz 10%, lc u. Cho t t theo thnh ng nghim 2ml H2SO4 . IV. PHN NG ADAMKIEVIE IV.2. Quan st: ng nghim 1: gia 2 lp cht lng xut hin 1 vng mu tm . - ng nghim 2: gia 2 lp cht lng xut hin 1 vng mu tm cam. IV.3. Gii thch - ng nghim 1: tryptophan c mt trong lng trng trng s tc dng vi axit glyoxylic (c mt trong axit axetic m c) to sn phm ngng kt lm thnh mt vng tm gia 2 dung dch lng. Trong phn ng ny H2SO4 c vai tr kh nc. -H Tryptophan CH2-CH -COOH NH2 NH + HOC-COOH 2

Axit glyoxylic - CH CH2-CH -COOH NH2 NH CH2-CH -COOH NH2 NH COOH Sn phm ngng kt giatrytophan vi axit glyoxylic c mu tm - H2O - ng nghim 2: tryptophan c mt trong lng trng trng s tc dng vi oxymetyl furfurol (c trong fructoz ca saccaroz) to sn phm ngng kt oxymetyl furfurol tryptophan lm thnh mt vng tm cam gia 2 dung dch lng. O H OH H OH CH2OH CH2OH -3H2O O -CH2OH HOCH H Fructoz Oxymetyl furfurol -H Tryptophan CH2-CH -COOH NH2 NH O -CH2OH HOCH H

Oxymetyl furfurol + 2 - CH CH2-CH -COOH NH2 NH CH2-CH -COOH NH2 NH Sn phm ngng kt gi oxymetyl furfurol vi trytophan c mu tm cam O H H CH2OH IV.4. Kt lun: y l phn ng c trng pht hin tryptophan c trong protein. V.1. Cch lm: Cho vo ng nghim 1ml dd protein trng, 2ml dd NaOH 10% d. un si 3 pht, lc u. Thm vi git (CH3COOH)2Pb 5%, h nng. V. PHN NG TO CH SUNFUA V.2. Quan st: Trong ng nghim xut hin nhng ht ta mu xm, en l lng trong dung dch. V.3. Gii thch: Trong dd protein trng c cha cc axit amin c S (xystein, xystin). Khi un vi kim d v mui ch s to kt ta sunfua ch l lng trong dung dch. HS- CH2 -CH -COOH NH2 + 2 NaOH HO- CH2 -CH -COOH NH2 + Na2S +H2O +NaOH d + 2 NaOH (CH3COOH)2Pb 2CH3COONa

+ Pb(OH)2 + NaOH d Pb(OH)2 + 2 NaOH Na2PbO2 + 2 H2O + NaOH d Na2S + Na2PbO2 + 2 H2O PbS + 4 NaOH V.4. Kt lun: y l phn ng c trng pht hin axit amin cha lu hunh c trong protein.