Инструментални методи-Скрипта-1 del bez UV VIS.pdf

1

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13

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Eabs, (xtrue)

(xi), :

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2.4.2.

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SD.

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(2)

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CV = [SD / ] 100%.

14

2.4.3.

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101 cm3.

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15

2.4.4.

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(Z = X+Y) (Z = X-Y): (3)

(Z = X Y) (Z = X/Y): (4)

, X X, Y

Y, Z

Z.

.

.

(ME)

(MI):

ME = 3.1 0.3 g

MI = 10.5 0.7 g

% = 100 ME / MI

,

4, .. (Z=X/Y).

:

X = 3.1; X = 0.3

16

Y = 10.5; Y = 0.7

% = Z = 100X/Y = 1003.1/10.5 = 29.5%

Z = Z [(X/X)2+(Y/Y)

2] = 29.5% [(0.3/3.1)

2+(0.7/10.5)

2] = 3.5%

, 29.5 3.5%.

2.4.5.

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12.13, 12.1

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(Z = X Y) (Z = X/Y),

(Z)

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12.312 (5 ) x 31.1 (3 ) = 383 (3 ).

(Z = X + Y) (Z = X - Y),

(Z) Z

(X Y) .

, 123.4567 ( "0.0001" ) + 0.31

( "0.01" ) = 123.77 (

"0.01" ). ,

5, 0,, 5

5, . 23.453 23.45; 23.455

23.46; 23.458 23.46.

17

2.4.6. :

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18

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2.4.7.

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Q-test:

XBAD , XNEXT

XBAD, XHIGH ,

XLOW . Q

Q-test

, :

19

Q-

(90% )

3 0.94

4 0.76

5 0.64

6 0.56

7 0.51 0.47

8 0.47 0.44

9 0.44 0.41

, 5

( 20, 22, 25, 50, 21), (

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, Q- 0.64, .. Q-

Q-test .

20

, ,

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21

(UV-VIS)

(UV)

X-, .

1nm 400nm.

UV

UV-A:

UV,

320 400 nm

UV-B:

UV

280 320 nm

UV-C:

UV ,

180 280 nm

UV-A UV-C: UV .

22

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80 1100nm (

), .

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Beer-Bougouer- - .

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24

()

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3.3. UV Vis

3

: (185-400nm), (400-700nm)

(700-1100nm).

185

900nm.

( ), ,

190 nm.

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) ,

150nm. .

,

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3300nm.

()

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""

.

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(Eelec),

(, ) :

Erot i Evib.

Eelec Erot Evib,

,

, ()

,

: charge transfer spectra.

Etot = Erot + + Evib Eelec

25

Eelec> Evib> Erot.

2. .

, .

Figure 2.Energy quantification of molecules. The diagram shows the electronic,

vibration and rotational energy state of the molecule.

.

: ,

S, (V) ,

(R)- ( -

), Eelec: Evib: erote 1000:50:1.

, ,

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:

- (

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- (

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26

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Kirchoff, Bunsen, Angstrom, , Michelson almer : "

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().

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3.

. .

Figure 3.Sample is translated in atomic conditionwith the use of the flame. Absorbed

lightness comes from the fixture

27

,

( )

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70 .

4.

Figure 4. Scheme of AAS

,

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. Kao

, ( , )

.

5.

Figure 5. Emission spectroscopy analysis

28

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0,0001%.

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(2100 ) N2O (2700 ).

3.6

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29

():

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6.

Figure 6. Scheme of spectrophotometar

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.

/ ( ) Vs. ( nm, m-1). , - T, "" ( ) , (I) (I0). , ( , ) " " .

- .

30

, : H, C, N, . , , (), , (L mol-1 cm-1), a . : - *n - *n - * - * d d

7.

Figure 7. Scheme of electronic crossings

--> * ,

M M

. O ,

, UV:

( ): max = 135nm

-

.

n *

31

n O, N, S, Cl *

180nm , 190 nm

220nm .

: max = 183nm; : max = 190 nm;

-: max = 210nm, 1-: max = 179nm.

n ( a * ).

() .

,

270-295 nm.

: e: max = 293nm, ).

*

,

170 nm.

: : max = 165nm

d - d

d ,

VIS a.

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C ),

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33

8.

Figure 8. Electronic Absorption bands for representative chromophores

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max

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34

pH :

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3.7.

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9.

Figure 9. Scheme of a simple beam, sequential mode spectrophotometer

10.

36

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0,01 nm.

11.

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x . Ho2O3,

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3.8.

-

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Lambert-Beer, , ,

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(L -1cm-1), .

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200.000).

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2.Ko

3.

4.

5.

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39

12.

Figure 12. Parts of a spectrophotometer

,

:

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IR

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40

4.

-

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(UV/VIS)

(100-380 nm) (380-800 nm)

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41

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42

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43

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2000 - 3000 oC.

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45

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AES [24].

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AES

[24].

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46

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(ICP).

AES

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AES

6000 10000

K () . ,

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(Ar Ar+ + e

-)

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- M

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47

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ICP-AES ICP-MS

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(iductively coupled plasmamass spectrometry, ICP-MS),

(multi-

element detector) ,

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48

). 100-1000

-

(iductively coupled plasma-atomic emission spectroscopy, ICP-

AES).

ICP-ES ICP-MS

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AES .

ICP-AES ,

2-3 % HNO3,

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(quartz torch),

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10 000 .

49

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,

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ICP

ICP ( )

, MS.

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.

50

. ICP-AES

51

,

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. 1, 2 , 3

, 4-

(2),

, .

3,

( )

(4).

.

ICP :

-

-

- .

52

ICP-MS

. ,

As, Cd, Cr, Cu, Fe, Mn Ni, Pb, Sn, V Zn. , ICP-

MS

, , ., ,

Pb, Cd, Zn, Cu Fe,

.

53

,

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.

(Scott 1992).

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( ,

).

54

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graphien .

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(Hight Pressure

Liquid Chromatography) ( ),

(High Performance Liquid

Chromatography HPLC).

11.1.

:

t (tR)

-

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,

:

, , .

55

(kA, kB) -

. :

M

MARA

)(

t

ttk

Rt

M

MBRB

)(

t

ttk

(tR)A (tR)B , tM e

a

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A

B

k

k

(RS) -

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BABA

ARBR 2)()(2sww

Z

ww

ttR

WA WB

, a .

(

),

.

-

. (N)

(dp),

(L) (F) mL/min.

:

56

2

16

w

tN R

tR e w .

(H), ,

.

(L) , .. H = L/N.

, (Skoog et al.

1994).

11.2.

.

.

, :

(), , , -

. ,

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.

: , , - .

11.3.1.

()

. -

(chemialy bonded phase chromatogaphy).

. -

57

, , ,

, ,

.

OH- , -

[8, 10].

, -Si-O-Si- .

OH- - -

:

Si O H + Si R

C H 3

C H 3

Si O Si R

C H 3

C H 3

Cl + H Cl

R (-C8H17) (-C18H37) .

,

, ,

, . ,

.

,

.

, -

(normal-phase HPLC) -

(reversed-phase HPLC). -

(, ),

(, i-),

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, (),

(, , )

.

58

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: )

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; )

(

) ( ).

A (+),

(. 2) .

, ,

:

() ()

() ()

59

S

P

C

CK

K

CP ;

CS

,

:

AS

APA

C

CK

BS

BPB

C

CK

A > ,

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(. 2). , KA

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, - , ,

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(, , .) .

60

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61

2.

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; ) )

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. :

, (Harris, 1995).

, ( 500 g/mol),

(,

),

( , ,

.). ,

,

. . 3

.

() ()

() ()

()

62

( ) .

, .

,

.

,

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.

( . ),

,

. ,

.

,

,

, .

3.

63

()

)

.

.

,

.

(

)

. ,

. ,

(Skoog et al. 1994).

11.3.2.

,

(

),

(, ,

) .

11.3.2.1.

(

)

.

. ,

. ,

64

,

.

,

.

, ,

.

11.3.2.2.

. : ,

, ,

.

, ,

. ,

. ,

,

: , ,

, , ,

, , , , , , ,

, , , ,

.

11.3.2.3.

(LC)

(0.10 0.25 mm)

,

, .

.

, ,

65

(.

4).

(UV).

4. LC

11.3.

, : ) ; )

; )

)

. ,

- : < K B < K C,

,

3 ,

. ,

.

)

, .

D

; A

< K B < K C < KD. D

. , ,

.

, , .

66

, , ,

D.

)

. ,

( K A < K B < K C) ,

.

.

, , : ,

, .

11.3.1. -

,,

HPLC,

High performance liquid chromatography. O

.

.

( ),

,

.

HPLC :

,

,

(),

,

,

,

67

(, 2005).

5.

- ( 4)

.

.

1 L.

, (Scott, 1992).

-

,

. , ,

,

.

. 40 MPa.

, , (

68

)

.

, :

, ..

;

, ..

.

() -

.. . ,

.

-

[9-11],

. HPLC

( , ,

). ,

, , .,

,

. , , .. (guard coulmns)

0,4 2 cm,

.

, .

,

,

.

,

.

69

,

. ,

(

) (Skoog et al. 1994).

( 10-9 10-5 g/L)

,

. ,

. ,

,

. ,

, . ,

,

.

-

(UV) - (UV-

VIS) , (

254 nm), , (diode-array

detector, DAD). ,

.

UV UV-VIS .

,

. ,

, ..

70

(, 2004).

DAD UV-Vis

.

UV-Vis .

UV

.

, .

.

() (n)

, , (2 - 1).

,

-

.

HPLC

. ,

, .

,

, . ,

,

.

,

.

, ,

.

=

2-

1

n

71

- ,

.

, ,

. ,

(Raymond Scott, 2003).

.

.

,

:

.

, ,

,

,

, .

, 1000

UV-.

( ),

, ,

. ,

UV (. ).

72

(, , ,

, , .).

Ag/AgCl ,

.

.

,

.

.

.

.

11.4.2.

(MS)

,

, , , ,

.

MS ,

/ (m/z) . MALDI-TOF

,

,

, .

.

(

) .

73

, , , . ,

( ).

, (Watson

Sparkman, 2007).

, ,

, .

, .. .

(Watson Sparkman, 2007).

. ,

.

, .

.

,

m/z ,

100 %.

,

, (Watson Sparkman,

2007).

MS :

, .

(Lavagnini et al., 2006).

-

(, , ,

74

). ,

:

,

.

(EI)

(CI). , ,

( ),

(APCI), (APPI),

(matrix-assisted laser desorption/ionization, MALDI,

/ ).

(EI)

(70 eV)

( 10-7 - 10-5 Torr) (Lavagnini et al., 2006).

. , ,

+,

:

- - = + + 2- = +

.

,

( ).

.

, :

(

75

,

),

( ,

) (, 2009).

,

[M+H]+ [M-H]- , (Lavagnini et al., 2006).

, ,

(PA) ,

.

189 kcal/mol (PAM),

CH5+ (PACH4 = 127 kcal/mol) H3O

+ (PA H2O = 165 kcal/mol),

NH4+ (PA NH3 = 205 kcal/mol).

,

. ,

- [AH]+, [H]+

PA .

(APCI)

,

,

. ,

0,1-1 Torr, ,

, ,

76

, (Lavagnini

et al., 2006).

APCI ,

.

(: corona discharge).

. ,

( 350-400 C), ,

(corona discharge)

. ,

.

. ,

,

.

( ) () .

, .. ()

.

, , ,

( N2) ,

(, 2009).

(ESI)

, .

ESI :

-

,

-

,

,

77

- ,

.

, ,

(

10-4 m) 2-5 kV, , ,

1-3 cm, , ESI

( 100-500 ; 5-10 cm) interface

. ,

,

.. Taylor .

, .

, .. ,

,

. .

.

, ,

.

.

()

, . ,

, ,

.

(Lavagnini et al., 2006).

(APPI)

,

. 10,6 eV

<

78

(Lavagnini et al., 2006). ,

, (IEM)

v = hv, :

IEM __ v

IE 10,6 eV,

r-.

APPI , LC/MS

IE 10,6 eV. IE > 10,6 eV,

(, 2009).

/ (MALDI)

,

( , 1:10 000).

,

,

,

, /

(Lavagnini et al., 2006). 11.5.

11.3.2. HPLC-DAD-MS

.

. ,

/ (.

79

)

, ,

.

HPLC,

UV

.

( , RID)

. HPLC-RID

,

. , RP (RP-

- )

. 2000 (Wey Ding, 2000)

, .

,

(Calull et al. 1999, Verette et al. 1995). , RID

(Akiyama et al. 1999, Vendrell et al. 2000)

(ED),

.

. XX

, .

80

.

6. HPLC (5 g/L)

(4.986 min), (6.190 min) (9.198 min)

(: = 75:25) 0,9 ml/min.

.

, .

.

. ,

, , ,

.

. ,

.

.

HPLC :

, ,

. /

: , , , .

, HPLC .

, .

81

, HPLC ,

.

. ,

,

.

(, ,

), (, ), ( )

pH . ,

(Morton et al. 1999).

j

. ,

(SEC), (AC),

- (SPE)

( C2 C18).

200-220 nm,

.

254 nm (,

) 280 nm ( ).

( )

.

, , ,

,

.

HPLC

(, ) (,

, .). . 7

220 nm.

82

. 7.

: C18 (10 cm x 4,6 mm I.D.), : (0,1

% : = 90:10) B (0,1 %

: = 25:75), ( 25 % B,

15 min 40 % B, 20 min 60 % B), 1,5 mL/min

1- , 2 , 3 - , 4 -

, 5 - C, 6

11.4. ALDI-TOF-MS ( /

- - )

MALDI (matrix-assisted laser desorption/ionization-

/ )

.

1987 Karas (Karas et al., 1987).

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11.4.1. MALDI-TOF-MS

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Tholey Heinzle, 2006; Carpentieri et al., 2007).

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109

et al. 2000, Servili et al. 2000, Cremer Eichner, 2000), -

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125

, ,

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Cl, F, Br, SH SeH -

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CSV - ;

ASV - a ;

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, . (2004).

, ,

, - , .

, .

, . (2009). , , -

, . , .

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