1

CC PHNG PHP PHN

TCH CNG C
2
M U
S pht trin ca cc ngnh vt l, ho l
S pht trin ca khoa hc k thut, cng ngh
Nhu cu phn tch nhanh, chnh xc, mi i
tng vi hm lng khc nhau
Pht trin cc phng php phn tch vt l v
ho l (phng php phn tch cng c)
Cc phng php phn tch cng c c xy
dng da trn mi tng quan gia cc tn hiu
o (vt l hoc ho l) v thnh phn ca h
nghin cu.
3
M U
Cc phng php phn tch quang hc
Quang ph nguyn t
Hp th (AAS)
Pht x (AES); ICP-OES
Hunh quang (AFS)
Quang ph phn t
Hp th (UV-Vis); Ph hng ngoi
Hunh quang (MFS)
Cc phng php phn tch in ha
o th
in lng
Von-ampe ha tan
Cc ph
Cc phng php phn tch sc k
Sc k kh (GC)
Sc k lng hiu nng cao (HPLC)
Sc k bn mng, sc k giy
Cc phng php tch
Chit lng lng (LLE)
Chit pha rn (SPE)
Chng ct

4
I CNG V PHNG PHP
PHN TCH QUANG PH
5
Phn loi cc PPPTQH
Cc phng php phn tch quang hc:
Ph hp th phn t (ph UV-VIS, ph hng ngoi)
Ph nguyn t (pht x nguyn t, hp th nguyn t)
Ph hunh quang (hunh quang phn t v nguyn t)
Cc phng php quang hc khc (o c, o khc
x, o quay cc)

6
u im ca cc PPPTQH
nhy tng Phn tch vt
Tnh chn lc cao
t tn thi gian
T ng ho
7
CC PHNG PHP PHN TCH
QUANG HC
Cc phng php phn tch quang hc da trn vic nghin
cu s tng tc ca bc x nh sng trn cht kho st
hoc s pht ra cc bc x nh sng di mt tc ng vt
l no .
Da vo nguyn tc ca phng php o ngi ta chia lm
2 nhm:
- Nhm 1: bao gm cc phng php da trn vic o
hp th ca dung dch mu phn tch hay hp th ca
m hi nguyn t to ra t mu phn tch. Ta c phng
php quang ph hp th phn t UV-Vis, Phng php
quang ph hp th nguyn t AAS.
- Nhm 2: bao gm cc phng php da trn vic o
cng chm bc x pht ra t mu phn tch hoc m
hi nguyn t do tc ng ca mt tc nhn vt l no .
Ta c phng php ph hunh quang phn t hoc ph pht
x nguyn t AES

8
NH SNG V BC X IN T
nh sng l mt phn trong ph cc bc x
in t.
Cc bc x in t gm: nh sng nhn thy,
tia t ngoi, tia hng ngoi, tia Rngen (tia
X), tia , sng radio,
9
10
NH SNG V BC X IN T (tip)
Bc x in t c bn cht hai mt: tnh sng v tnh ht
Tnh cht sng: Cc bc x in t l nhng dao ng c hai
thnh phn in trng v t trng lan truyn theo mt phng,
vi vn tc bng vn tc nh sng c (c=3.10
10
cm/s). Cc thnh
phn in trng v t trng cu sng in t nm trong hai
mt phng vung gc vi nhau v vung gc vi phng truyn
ca bc x.
Mt phng dao ng trong in trng
Mt phng dao ng trong t trng
Hnh 1.1. Thnh phn in trng v t trng ca bc x in t
Phng
truyn
11
NH SNG V BC X IN T (tip)
Cc dao ng c bin bin thin theo thi gian
khi lan truyn
Cc dao ng c c trng bng bc sng
hay tn s v
Thnh phn vect in trng ca bc x in t
tng tc vi cc nguyn t hoc phn t gy nn
hiu ng hp th v to ra ph hp th nguyn t
hoc phn t.
12
NH SNG V BC X IN T (tip)
Tnh cht ht
Bc x in t l cc ht mang nng lng c gi l
photon lan truyn vi vn tc nh sng.
Cc bc x in t c bc sng khc nhau c nng
lng khc nhau.
gy ra hiu ng hp th bc x in t, nng lng
ca bc x phi bng hiu mc nng lng AE tng
ng vi cc trng thi nng lng ca nguyn t hoc
phn t. Ngha l bc sng ca bc x in t phi
ph hp vi phng trnh:
AE = hc/ = hv (1.1)
h = 6.63.10
-34
j.s c: vn tc nh sng, cm/s
Phng trnh (1.1) th hin s thng nht bn cht sng
v ht ca bc x in t.
13
Bc sng : qung ng m bc x i c
trong 1 dao ng. (m, m, nm, )

Tn s v: s giao ng m bc x in t thc
hin c trong 1 s, th nguyn (s
-1
), n v Hz,
kHz, MHz.
v = c/ (1.2)
S sng v : v = 1/ (cm
-1
) (1.3)
NH SNG V BC X IN T (tip)
CC I LNG C TRNG CHO BC X NH
SNG
14


Khng kh Khng kh Thu tinh
v = 6,0.10
14
Hz
= 500 nm
v = 6,0.10
14
Hz
= 330 nm
v = 6,0.10
14
Hz
= 500 nm
15
NH SNG TRONG PH SNG IN T, THANG
SNG IN T V CC PHNG PHP PH
min
nhn
thy
Tia
gn xa
min t ngoi
xa gn
min hng ngoi
Sng
viba
10
3
10
12
10
17
10
20
Hz
10
12

10
14
4.10
14
7.10
14
15.10
14

10
17
Hz
Sng radio
nh sng quang hc
Tia Rngen
16
NH SNG V BC X IN T (tip)
Tu thuc vo bn cht ca bc x in t tng
tc vi nguyn t, phn t m ta c cc phng
php phn tch ph khc nhau.
Min sng radio, vi sng: ph cng hng t
Min tia Rngen, tia : phn tch ph Rngen, ph
tia
Min nh sng quang hc: ph hp th phn t ( ph
nhn thy (VIS), ph t ngoi (UV), ph hng ngoi
(IR)
Trong min ph quang hc c cc phng php ph
hp th, ph pht x, ph hunh quang.
17
S HP TH NH SNG V MU SC
Theo thuyt lng t, mi ht s cp (phn t, nguyn t,
ion) c mt h thng duy nht cc trng thi nng lng.
Trng thi nng lng thp nht gi l trng thi c bn.
S hp th phton nh sng bi ht s cp ch c th xy
ra khi phton mang nng lng ng bng hiu cc
mc nng lng ca 1 trng thi nng lng cao hn v
trng thi c bn.
Trng thi nng lng cao hn gi l trng thi kch thch.
M + hv M*
Sau mt thi gian rt ngn (10
-6
10
-9
s), tiu phn trng
thi kch thch s phc hi tr v trng thi c bn ban u
c nng lng thp hn, truyn nng lng tha cho cc
nguyn t v phn t ca mi trng xung quanh lm cho
mi trng nng ln mt t.
M* M + nhit (phc hi khng bc x)
18
S HP TH NH SNG V MU SC
S phc hi c th thc hin thng qua s
phn hu quang ho ca M* thnh cht
mi hoc bng s pht x hunh quang
(phc hi c bc x).
c tnh hp th ca mt cht c m t
bng ph hp th quang, l ng biu
din hp th A theo hay v
19
MU SC
nh sng c chia thnh 3 vng:
Vng t ngoi: 185-400 nm
Vng kh kin: 400-760 nm
Vng hng ngoi: 760 1000 nm

Vng kh kin c nhiu mu, mi mu ng vi mt chm
cc tia c bc sng gn nhau:

: 760 - 630 nm Lam: 500 450 nm
Cam: 630 600 nm Chm: 450 430 nm
Vng: 600 570 nm Tm: 430 400 nm
Lc: 570 500 nm
20
MU SC
Mu ph nhau: Mt vt c mu lam hp th mnh cc tia
mu vng v ngc li vt c mu vng hp th mnh cc
tia mu lam. Hai mu vng v lam hnh thnh mt cp
mu ph nhau hay b sung cho nhau.
Cc cp mu ph nhau:
Lc vng - Tm
Vng - Lam
Da cam - lam lc
- Lc lam
ta - Lc
nh lng mt dung dch mu bng phng php
o hp th, ngi ta dng knh lc c mu ph vi mu
dung dch cn o.
21
22
23
S HP TH CA NGUYN T
Khi chiu mt chm tia t ngoi v kh kin qua
mt mi trng hi nguyn t th ch c mt s t bc
x c tn s xc nh b hp th v ph hp th thu
c gm mt s vch hp th rt hp (gi l ph
vch).
200 300 400 500 600

4,0

3,0

2,0

1,0

0,0

Nng
lng
electron
volts
285
nm
330
nm
590
nm

hp
th
3s
3p
4p
5p
Ph hp th ca hi Na Gin mc nng lng
24
S HP TH CA NGUYN T
25
1. Trng thi nng lng ca phn t
E
tng
= E
t
+ E
e
+ E
v
+ E
r

E
t
: nng lng ca chuyn ng tnh tin
E
e
: nng lng ca in t
E
v
: nng lng ca chuyn ng dao ng
E
r
: nng lng ca chuyn ng quay
Trong : E
e
>> E
v
>> E
r
;
(E
e
~ 60 150, E
v
~ 1 10, E
r
~ 0,01 0,1 kcal/mol)

M u
PH PHN T
26
1. Trng thi nng lng ca phn t (tip)

S hp th bc x hng ngoi ca phn t lm thay i
mc nng lng ca chuyn ng dao ng v chuyn
ng quay ca phn t (E
v
v E
r
) l ngun gc ph
hng ngoi ph hng ngoi c gi l ph dao ng.
S hp th bc x vng t ngoi, kh kin ca phn t
lm thay i mc nng lng ca in t ha tr trong
phn t
Ph hunh quang phn t l s pht ra bc x ca cc
phn t c kch thch bng cc bc x nh sng. y
l phng php ph pht x phn t.
Ph phn t l ph gii, ph lin tc.
PH PHN T
27
S HP TH NNG LNG CA PHN T
Hunh
quang cng
hng
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
4
3
2
1
0
E
0

E
1

E
2

(a) Hp th phn t (b) Phc hi khng bc x (c) Hunh quang
Nng
lng
IR VIS UV
E
1

E
2

E
0

E
0

E
1

E
2

28
S HP TH NNG LNG CA PHN T
E
0
, E
1
, E
2
: cc mc nng lng in t ca trng thi c
bn v cc trng thi kch thch
ng vi mt mc nng lng in t li c mt s mc
nng lng dao ng.
ng vi mi mc dao ng li c mt s mc nng lng
ca chuyn ng quay.
Hnh (a) ch ra: s hp th bc x t ngoi (UV) c nng
lng ln dn n chuyn mc nng lng ln E
2
, VIS
E
1
; IR thay i E
v
, phn t vn trng thi nng lng
c bn.
Ph hp th phn t c dng l gii ph (ph m)
S phc hi nng lng do chuyn nng lng ca phn
t b kch thch cho dung mi (khng bc x) hoc pht ra
hunh quang.
29
CC BC CHUYN IN T V PH UV-VIS
TRONG PHN T
30
PH HP TH CA PHN T
31
2. S hp th BXT v trng thi nng lng ca phn t
- iu kin bnh thng phn t trng thi c bn, c nng
lng thp ( E
o
)
- Khi phn t nhn nng lng ( bc x, nhit..), phn t chuyn ln
trng thi nng lng cao hn, gi l trng thi kch thch (E*)
E*
tng
= E*
e
+ E*
v
+ E*
r

AE
t
= E
t
* - E
o

= (E*
e
- E
o
) + (E*
v
- E
o
v
) + (E*
r
E
o
r
)
= AE
e
+ AE
v
+ AE
r
S hp th nng lng bc x tng ng vi bc chuyn mc
nng lng no th s to ra loi ph hp th phn t tng ng:
AE
e
: Ph hp th phn t UV-VIS
AE
v
: Ph dao ng phn t
AE
r
: Ph quay phn t
32
2.1. PHNG PHP PH HP TH T NGOI
KH KIN
2.1.1. CC VNG PH
Trong ph cc bc x in t, cc tia t ngoi v kh
kin ch chim mt phn rt hp:
bc sng
Tia X = 0,1 50 nm
Tia UV xa = 50 200 nm
Tia UV gn = 200 400 nm
Tia Vis = 400 800 nm
33
Tia UV xa: Bc x ny t c dng v:
C nng lng kh cao lm ph v lin
kt trong cc phn t
B hu ht cc dung mi hp th mnh, nn
c th lm ho hi dung mi
B thch anh (quartz) hp th
xi trong khng kh cng hp th mnh bc
x c = 180 nm
34
Tia UV gn v Vis: thng c dng v:
Lm thay i nng lng in t trong
phn t
Thch anh (quartz) khng hp th trong
min ny.

Cc my o hp th UV-Vis thng o
c cc bc x c bc sng = 190
1000 nm, dng cuvet thch anh. i vi tia
Vis c th dng cuvet thu tinh hoc cuvet
nha.
35
Cc in t lin kt:
Trong Hydrocacbon no:
in t o c th c trong lin kt C-C hoc C-H
in t o ln c trng thi kch thch o* cn c
nng lng E ln v khong cch gia cc mc o v
o* ln.
Trong Hydrocacbon cha no:
in t t c trong lin kt i v lin kt ba
C = O, C = C , -C C-
in t t ln trng thi kch thch t* cn c nng
lng E nh hn v hai mc ny gn nhau hn.
Nu phn t cng c nhiu dy ni i th s hp th
cng chuyn v pha bc x c bc sng di
Nu phn t c ni i lin hp
C = C C = C th u tin hp th bc x c bc
sng di hn (bc x kh kin, vng ).
2.1.2. Cc in t tham gia vo s hp th
36
Cc in t phi lin kt:

in t n: l i in t t do khng
tham gia lin kt (vd: O, N, Cl). C hai
loi in t t do n
s
v n
p
c th chuyn
ln mc nng lng t*, trong cc
in t n
s
c mc nng lng nh hn n
p
S hp th thng trong min t 150
250 nm v gi tr thp
37

M t cc mc nng lng:


S hp th UV-Vis ca cht hu c i hi nng
lng hp th tng ng vi bc nhy t orbital
c in t ln orbital trng. S chuyn mc nng
lng t n t* cn nng lng thp nhng xc
sut xy ra li thp hn s chuyn t t*.
o
*

t
*

n
p

n
s

t
o
Phn lin kt
Phn lin kt
Khng lin kt
Khng lin kt
Lin kt
Lin kt
38

Bc chuyn t o o* ,
t c s dng
cc i hp th < 150 nm
Nng lng qu ln
Loi hp th ny tng ng vi ph v lin kt
C-C, C-H, C-O, C-X

39
40
41
42
43
nh lut hp th nh sng (L Bougeur-
Lambert-Beer)
Khi chiu 1 chm bc x n sc c cng l I
0
i
qua mt lp dung dch cht mu, cng chm bc
x i ra khi dung dch l I
t

Gia I
t
v I
0
lin h vi nhau theo biu thc:
I = I
0.
10
-K

Trong K ph thuc vo b dy lp dung dch ( l);
nng dung dch (C ) v bn cht ca dung dch (c)
I = I
0.
10
-clC

Nh vy I
0
/I = 10
clC
=> lg I
0
/I = clC

= A ( hp th
quang)
I/I
0
: truyn quang (T) => A = log 1/T
44
Cc tnh cht ca hp th
T biu thc nh lut hp th nh sng ta c
A = clC
c l i lng ph thuc bn cht ca cht hp th v
bc sng ca nh sng chiu vo. Nu C tnh bng
mol/L th c gi l h s hp th phn t gam.
H s hp th phn t gam c trng cho nhy ca
phng php phn tch
A l hm theo cc bin l C, l, v
ng biu din s ph thuc ca A ~ C l ng
thng: c dng trong phn tch nh lng
ng biu din s ph thuc A ~ l mt ng
phc tp c trng cho cu to ca cht phn tch gi l
ph hp th: c dng trong phn tch nh tnh v cu
to cht
hp th c tnh cht cng tnh
45
2.1.3. Cc nhm chc ho hc tham gia
vo s hp th nh sng
Nhm mang mu (chromophore)
Cc cht c mang cc nhm ny ch yu hp th
cc bc x c bc sng >200 nm.
Cc nhm
c s chuyn n t
*
, Hp th cc bc x c bc
sng = 300 305 nm.
Cc nhm
c s chuyn in t t t
*
nn ch yu hp th
cc bc x c bc sng > 200 nm.
46
Nhm tng mu (auxochrome):
Gm cc halogenua (Cl
-
, Br
-
, I
-
); cc alkyl (-
CH
3
, -C
2
H
5
); cc amin (-NH
2
, -NH-) v nhm alcol
(-OH),
Cc nhm ny hp th bc x c < 200 nm.
Kt hp cc nhm ny vi nhm mang mu lm cho
s hp th ca hp cht chuyn v pha c di hn.
Nhm lm cng hp th tng ln gi l nhm
gy hiu ng Hyperchromic
Nhm lm cng hp th gim i gi l nhm
gy hiu ng Hyphochromic
47
2.1.4. CC YU T NH HNG LN
HP TH

2.1.4.1. Cc yu t thuc v cu trc phn t

Hiu ng cm ng
Hiu ng khng gian
48
2.1.4.2. Cc yu t thuc v mi trng
a. Dung mi:
Dung mi c th hp th bc x UV-Vis cn phi m rng
khe sng tng cng sng. Khi tia sng km n sc
i nhy ca php o gim.
Mi dung mi dng cho bc x UV-Vis ch dng trong khong
bc x c bc sng nht nh.
V d:
Dung mi nc: dng cc bc x c > 205 nm
Metanol, cyclohexan, acetonitril v isopropanol dng
>210 nm
Tetrahydrofuran dng > 220 nm
Dichloromethan dng > 235 nm
Chloroform dng > 245 nm
Aceton dng > 330 nm
Ch : Khi dung mi hp th bc x mnh th php o mc sai
s ln. La chn dung mi cn quan tm n tan ca cht
cn o hp th trong dung mi.
49
b. Tng tc gia hai lng cc (Dipole-Dipole)
Khi dng cc dung mi phn cc nh nc,
acetonitril, metanol, nu cht phn tch cng lng
cc th c tng tc lng cc gy tc dng:
Khong cch dch chuyn t t* ngn li,
max
tng
ln (Hiu ng bathocromic: cc i hp th chuyn
v pha bc sng di)
Khong cch dch chuyn n t* di ra,
max
gim
(Hiu ng hypsochromic: cc i hp th chuyn v
bc sng ngn).
Vy khi thay i dung mi th
max
thay i.
50
c. Lin kt hydro
Cc dung mi c lk hydro nh ethanol c th tc ng
ln in t n trong phn t. Ngc li cc dung mi c
in t t do n li nh hng ln cc cht phn tch c
mang lk hydro.
Tc ng: lm khong cch dch chuyn n t* di ra
( hiu ng hypsochromic)
d. pH
Nu cht phn tch c tnh cht trao i proton (tnh
axit-baz), v d:
Hp th bc x
c = 285 nm
Hp th bc x
c = 293 nm
51
2.1.5. CC NG DNG
2.1.5.1. nh tnh v th tinh khit
Da vo
max:
V d:
Vitamin B
12
c 3 cc i hp th cc bc sng:
278 nm 1 nm
361 nm 1 nm
548 nm 1 nm
Vitamin B
2
c bn hp th cc i cc bc sng:
223 nm, 267 nm, 375 nm v 444 nm.
Da vo t s gia cc gi tr hp th cc i hoc cc tiu
nh tnh
V d:
Vi vitamin B
12
, da vo A
278
/A
361
= 0,57
hoc A
548
/A
361
= 0,3
Ngy nay ngi ta khng dng phng php ny nh tnh
na v quang ph UV-Vis cho qu t thng tin, trong khi
quang ph R li c nhiu thng tin hn v c trng hn.
52
5. iu kin ng dng ca nh lut Lambert Beer:

Chm tia sng phi n sc
Dung dch phi long
Dung dch phi trong sut (tr chun o quang)
Cht phn tch phi bn trong dung dch v bn
di tc dng ca tia UV-Vis
53
6. S sai lch i vi nh lut Lambert Beer
Sai lch do my (do b phn n sc, b phn khuch
i km) nh t bo quang in, nhn quang qu gi-
nhy km, h knh quang hc khng trong sut
trong di sng o.
Sai lch do ho hc:
Do s phn ly ca phn t cht phn tch
Do s to thnh dimer, trimer: sn phm trng hp
ny c hp th khc dng n phn t.
Do phn ng ca cc cht l c mt trong dung
dch phn tch: loi tr nh hng ny, ngi
ta dng mu trng (Mu blank, c thnh phn nh
dung dch, ch tr cht cn phn tch).
54
Cc k thut nh lng
a. o ph trc tip
b. Phng php so snh
c. Phng php thm
d. Phng php ng chun
e. Phng php thm ng chun
f. Phng php nh lng hn hp
g. Phng php chun o quang

55
ng dng ca php o ph UV-VIS trong ng/cu
s to phc
1. Xc nh thnh phn phc: thng s dng 3 phng php
sau:
a. Phng php bin i lin tc 1 hp phn
a
b
c
d
1
2
56
b. Phng php dy ng phn t:
Nhiu phc bn c thnh phn
khc nhau
Mt loi phc c thnh
phn xc nh
1
2
3
X R
X R
57
c. Phng php logarit gii hn
Phng php ny ch p dng i vi nhng phc rt
km bn.
Gi s c phn ng to phc: X + nR = XRn
K
b
= [XRn]/[X].[R]
n =>
[XRn] = K
b
. [X].[R]
n
[XRn] t l vi hp th A v v phc km bn nn [X] ~ C
x
v [R] ~ C
R

=> log A = log K
b
+ logC
X
+ nlogC
R
= K + nlog C
R
lp dy th nghim, v v thi logA ~ C
R
ta xc nh c n
tgo = n
log A
logC
R

58
2. Xc nh hng s phn ly ca phc km bn
a. Phng php ng chun: gi s c phc XR vi cn bng
phn ly : XR = X + R
=> K
f
= [X].[R]/[XR]
Phn ng to phc gia X v thuc th HR pH xc nh nh
sau: X + HR = XR + H
+

Hng s cn bng ca phn ng to phc ny l:
K = [XR].[H
+
]/[X].[HR]
HR c cn bng phn ly: HR = H
+
+ R
-

=> k
1
= [H
+
] [R
-
]/[HR]
=> k
1
/K = [X].[R]/[XR] = K
f

Nh vy tnh K
f
phi tnh c hng s cn bng K ca
phn ng to phc:
- Lp th chun A ~ C
X
=> [XR]
- [X] = C
x
- [XR]; [HR] = C
HR
- [XR]
T ta tnh c K
f
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b. Phng php dy ng phn t
Gi s c phc XR vi cn bng phn ly nh sau: XR = X + R
K
f
= [X].[R]/[XR]
Goi o l phn ly: K
f
= o
2
C
2
/(1- o)C
= o
2
C
Da vo th dy ng phn t ta c
o = (A
0
A
1
)/A
0
=> K
f

A
1

A
0

X
R
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2.1.6. MY QUANG PH UV-Vis
Ngun sng:
n Vonfram: o vng Vis vi > 320 nm
n Deuteri (D
2
) o vng UV vi < 350 nm (khi
o dng cuvet thch anh)
Khe sng: to ra mt chm tia sng c cng ln
chiu vo b n sc
B n sc:
Dng knh lc
Dng lng knh
Dng cch t
Detector: Nhn quang, t bo quang in, diode array
X l tn hiu: Ni my tnh
Cc loi my: Mt chm tia, Hai chm tia
61
S khi ca mt my quang ph hp th phn t
mt ng truyn o ti mt bc sng la chn
62
S khi ca mt my quang ph hp th phn t
hai ng truyn c kh nng qut ph
63
S khi ca mt my quang ph hp th phn t
mng diode
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65
66
67
68
CCH T NHIU X
69