Pesticides Identification at the Residue Level - ACS Publications

9 Applications of Combined Gas Chromatography-Mass Spectrometry to Pesticide Residue Identifications

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F R A N C I S J . BIROS Perrine Primate Research Branch, D i v i s i o n of Pesticide Chemistry a n d Toxicology, F o o d and D r u g Administration, U. S. D e p t . of Health, Education, and Welfare, Perrine, F l a . 33157

Several

individual

and multiresidue

analytical

available

for gas chromatographic-mass

firmation

and identification

examples

considered

intact and derivatized

phenolic

phenol and 1-naphthyl secticide

metabolic

-diethyl O-methyl

of pesticide

in this report

residues

chloroacetate,

pesticides

type, phenoxyalkanoic

acid herbicide

studies

rinated

involving

biphenyl

methodology

2,4-D

residues. directly

and environmental

nique.

Evaluation

identity

definitive

method-

and

polychlo-

residue column

analytical technology human,

by the combined

of mass spectral

fragmentation

and conclusive

confirmation

of unknown

cyclodiene

and

to the analysis of

substrates

as well as characterization

metabolites

2,4,5-T,

Conventional

applicable

animal, provides

and

phosphate,

and

exposure

inO,O-

S-methyl

O-methyl

of the DDT

and gas chromatographic

are, in general,

including

O,O-diethyl

O,O-diethyl

of

pentachloro-

organophosphorus

several organochlorine ology

Specific

the analysis

such as

products

phosphorothionate, and

are con-

residues.

include

and hydrolytic

phosphorothiolate,

methods

spectrometric

of residues

tech-

pathways of and

residue their

structure.

T T n e q u i v o c a l i d e n t i f i c a t i o n of p e s t i c i d e s , p e s t i c i d e m e t a b o l i t e s , a n d ^ other c h e m i c a l e n v i r o n m e n t a l p o l l u t a n t s i n m o s t cases r e q u i r e s m o r e evidence than can be p r o v i d e d b y a single chromatographic method. Seve r a l a p p r o a c h e s m a y b e u t i l i z e d to secure firm p r o o f of i d e n t i t y . F o r e x a m p l e , s u p p o r t i v e c h r o m a t o g r a p h i c d a t a s u c h as r e l a t i v e r e t e n t i o n 132

Biros; Pesticides Identification Advances in Chemistry; American Chemical Society: Washington, DC, 1971.

9.

BIROS

Gas Chromatography—Mass

133

Spectrometry

values o n t w o gas c h r o m a t o g r a p h i c c o l u m n s of d i f f e r i n g characteristics m a y b e o b t a i n e d . O f f u r t h e r v a l u e is i n f o r m a t i o n d e r i v e d b y means of t h i n - l a y e r c h r o m a t o g r a p h y (R

f

v a l u e s ) , the solvent p a r t i t i o n i n g c h a r a c -

teristics of the r e s i d u e of interest ( p - v a l u e s ) , a n d the response b e h a v i o r o n several m o r e o r less specific gas c h r o m a t o g r a p h i c detectors.

In addi-

t i o n , s p e c t r o m e t r i c t e c h n i q u e s s u c h as i n f r a r e d a n d u l t r a v i o l e t spectrop h o t o m e t r y m a y b e a p p l i e d to p r o v i d e e v e n less a m b i g u o u s p r o o f

of

r e s i d u e s t r u c t u r a l i d e n t i t y . T h e v a l u e of mass s p e c t r o m e t r y as a t o o l for p r o v i d i n g t h e s t r u c t u r a l i d e n t i t y of c o m p l e x o r g a n i c m o l e c u l e s has b e c o m e w e l l r e c o g n i z e d i n recent years. M a s s spectra f u r n i s h i n f o r m a t i o n Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

c o n c e r n i n g the s t r u c t u r a l a r r a n g e m e n t of atoms w i t h i n a m o l e c u l e o n the basis of the m o d e of f r a g m e n t a t i o n of the c o m p o u n d as a r a d i c a l i o n , usually produced b y electron bombardment.

F r a g m e n t a t i o n patterns r e -

s u l t i n g f r o m b o n d fission a n d r e a r r a n g e m e n t of atoms are h i g h l y d i a g n o s t i c a n d c h a r a c t e r i s t i c of the o r i g i n a l m o l e c u l a r structure. I n t e r p r e t a t i o n s are u s u a l l y m a d e o n the basis of e m p i r i c a l correlations, c o m p a r i s o n w i t h the mass spectra of s t a n d a r d m a t e r i a l s , or d e t a i l e d studies of i o n d e c o m p o sition mechanisms.

T h e theories a n d p r i n c i p l e s g o v e r n i n g the i n t e r p r e -

t a t i o n of mass spectra h a v e b e e n discussed i n m a n y reference texts a n d comprehensive reviews ( J , 2 ) . T h e p o t e n t i a l u t i l i t y of mass s p e c t r o m e t r y i n e v a l u a t i n g the m e t a b o l i c p a t h w a y s of p e s t i c i d a l c h e m i c a l s b y p r o v i d i n g the s t r u c t u r a l i d e n t i t y of metabolites w a s stated as e a r l y as 1962 i n a r e v i e w b y G u n t h e r M o r e r e c e n t l y , several r e v i e w s (4, 5, 6)

(3).

h a v e c o n s i d e r e d the r o l e of mass

s p e c t r o m e t r y i n c h e m i c a l s t r u c t u r e evaluations w i t h s p e c i a l reference to p e s t i c i d e r e s i d u e analysis. C o r r e l a t i o n s of mass s p e c t r a l f r a g m e n t a t i o n p a t h w a y s w i t h the s t r u c ture of pesticides are r e q u i r e d to o b t a i n b a c k g r o u n d i n f o r m a t i o n u s e f u l i n the i n t e r p r e t a t i o n of the mass spectra of u n k n o w n p e s t i c i d e m e t a b o l i t e s a n d other c o n v e r s i o n p r o d u c t s . F o r t u n a t e l y , the c h e m i c a l m o d e s of f r a g m e n t a t i o n of a l a r g e n u m b e r of p e s t i c i d a l c o m p o u n d s h a v e b e e n determined

(7).

T h i s i n f o r m a t i o n is p a r t i c u l a r l y u s e f u l i n those instances

w h e n o n l y m i c r o g r a m q u a n t i t i e s of m a t e r i a l s are a v a i l a b l e for c h a r a c t e r i z a t i o n ; mass s p e c t r o m e t r y offers one of the best i n s t r u m e n t a l a p proaches i n v i e w of the l o w s e n s i t i v i t y of s u c h c o m p l e m e n t a r y t e c h n i q u e s as n u c l e a r m a g n e t i c resonance spectrometry.

T h e investigator may then

r e l y o n this single source of i n f o r m a t i o n for a l l the s t r u c t u r a l d a t a . A n a d d i t i o n a l a d v a n t a g e of the mass s p e c t r o m e t r i c a p p r o a c h is the a v a i l a b i l i t y of c o m b i n e d gas c h r o m a t o g r a p h - m a s s spectrometer instruments w h i c h p e r m i t h i g h sensitivity analyses of m u l t i c o m p o n e n t m i x t u r e s . M a s s s p e c t r o m e t r y , of course, has b e e n w i d e l y e m p l o y e d as a n i n d i v i d u a l t e c h n i q u e for analysis of residues a n d metabolites i s o l a t e d b y c o n v e n t i o n a l separation t e c h n i q u e s s u c h as t h i n - l a y e r , l i q u i d , p a p e r , g a s - l i q u i d , a n d


134

PESTICIDES

column chromatography.

IDENTIFICATION

I n these instances, s p e c i a l i z e d t e c h n i q u e s are

e m p l o y e d f o r the efficient t r a p p i n g a n d c o l l e c t i o n of i n d i v i d u a l gas c h r o m a t o g r a p h i c peaks or t h i n - l a y e r c h r o m a t o g r a p h i c spots for

subsequent

mass s p e c t r a l analysis b y s t a n d a r d s a m p l e v a c u u m i n t r o d u c t i o n m e t h o d s . R e c e n t reports h a v e f o c u s e d a t t e n t i o n o n the n e e d for c o n f i r m i n g p e s t i c i d e residues, p a r t i c u l a r l y those i s o l a t e d f r o m e n v i r o n m e n t a l samples (8, 9 ) , a n d f o r c h e m i c a l i d e n t i t y i n u l t r a m i c r o a n a l y s i s i n g e n e r a l . O f t h e s p e c t r o m e t r i c t e c h n i q u e s a v a i l a b l e for c o n f i r m a t i o n of residues, the c o m b i n e d gas c h r o m a t o g r a p h i c - m a s s s p e c t r o m e t r i c a p p r o a c h affords

many

advantages i n c l u d i n g r e l a t i v e l y h i g h s e n s i t i v i t y , e l i m i n a t i o n of the necesDownloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

sity for i s o l a t i n g m i n u t e q u a n t i t i e s of p u r e samples w i t h s t a n d a r d c h r o m a t o g r a p h i c m e t h o d s , a n d a greater c e r t a i n t y i n i d e n t i f i c a t i o n of a n e l u t e d c o m p o n e n t t h a n that a c h i e v e d b y c o n v e n t i o n a l detector response c h a r a c teristics a n d gas c h r o m a t o g r a p h i c r e t e n t i o n times alone.

B e c a u s e of the

definitive s t r u c t u r a l i n f o r m a t i o n o b t a i n e d b y mass s p e c t r o m e t r y , p r o p e r use of the t e c h n i q u e w o u l d o b v i a t e t h e necessity for the a p p l i c a t i o n of t w o or m o r e different c a r e f u l l y selected t e c h n i q u e s for c h a r a c t e r i z i n g p e s t i c i d e residues

(8).

A n a d d i t i o n a l o b v i o u s a d v a n t a g e w o u l d b e the c h a r a c t e r i z a t i o n a n d i d e n t i f i c a t i o n of f r e q u e n t l y e n c o u n t e r e d , u n k n o w n c o m p o n e n t s of extracts, w h i c h m a y represent p e s t i c i d a l m e t a b o l i t e s , p h o t o c h e m i c a l a n d

other

" w e a t h e r e d " residues, n o n p e s t i c i d a l c h e m i c a l p o l l u t a n t s , or co-extractive interferences w h o s e i d e n t i t y m a y be r e q u i r e d . T h i s r e p o r t presents a d i s c u s s i o n of recent a p p l i c a t i o n s of c o m b i n e d gas c h r o m a t o g r a p h y - m a s s s p e c t r o m e t r y to analysis of p e s t i c i d e residues isolated from h u m a n , animal, a n d environmental media. Emphasis w i l l be p l a c e d o n a n a l y t i c a l t e c h n i q u e s , r e l a t e d gas c h r o m a t o g r a p h i c c o l u m n technology,

a n d analysis of f r a g m e n t a t i o n p a t h w a y s p e r t i n e n t to

the

i d e n t i f i c a t i o n of p e s t i c i d e residues.

Experimental T h e mass s p e c t r o m e t r i c analyses i n i t i a l l y r e p o r t e d i n this c o m m u n i c a t i o n w e r e p e r f o r m e d w i t h a d o u b l e f o c u s i n g l o w r e s o l u t i o n mass spect r o m e t e r ; M o d e l 270, P e r k i n - E l m e r C o r p . , N o r w a l k , C o n n . , c o u p l e d t h r o u g h a W a t s o n - B i e m a n n t y p e of m o l e c u l a r separator w i t h a gas c h r o m a t o g r a p h i c system. ( C o m m e r c i a l sources a n d t r a d e names are p r o v i d e d for i d e n t i f i c a t i o n o n l y . T h e i r m e n t i o n does not constitute endorsem e n t b y the P u b l i c H e a l t h S e r v i c e or b y the U . S. D e p t . of H e a l t h , Education, and Welfare.) C o i l e d glass gas c h r o m a t o g r a p h i c c o l u m n s w e r e e m p l o y e d except w h e r e o t h e r w i s e i n d i c a t e d . P r o g r a m m e d t e m p e r a t u r e analyses w e r e m a d e w i t h the i n i t i a l a n d final o v e n c o n d i t i o n s as w e l l as t h e p r o g r a m rate g i v e n i n the discussion. T h e m o l e c u l a r separator a n d gas i n l e t temperatures w e r e m a i n t a i n e d at 200° a n d 2 1 0 ° C , r e s p e c t i v e l y . A l l mass spectra w e r e


9.

BIROS


135

Spectrometry

r e c o r d e d at 80 e V e l e c t r o n e n e r g y w i t h 2400 V a c c e l e r a t i n g v o l t a g e ; t h e filament e m i s s i o n c u r r e n t w a s 100 /*a. C h r o m a t o g r a m s w e r e r e c o r d e d f r o m the t o t a l i o n c u r r e n t m o n i t o r l o c a t e d b e t w e e n t h e electrostatic a n d m a g n e t i c a n a l y z e r sectors. H e l i u m c a r r i e r gas w a s a p p r o x i m a t e l y 10 m l p e r m i n for the p a c k e d c h r o m a t o g r a p h i c c o l u m n s a n d a p p r o x i m a t e l y 2 m l p e r m i n for the c a p i l l a r y c o l u m n s . Injector t e m p e r a t u r e w a s m a i n t a i n e d at 165 ° C . M a s s s p e c t r a w e r e s c a n n e d m a g n e t i c a l l y over t h e r a n g e of interest at a rate of either 3 or 10 sec p e r decade. T h e c o m b i n e d gas c h r o m a t o g r a p h - m a s s spectrometer system has b e e n d e s c r i b e d i n m o r e d e t a i l elsewhere ( 1 0 ) .


Phenolic Residues C o n f i r m a t i o n of the i d e n t i t y of p e n t a c h l o r o p h e n o l ( P C P ) i n samples of h u m a n b l o o d , u r i n e , tissue, c l o t h i n g , a n d b e d d i n g m a t e r i a l s has b e e n reported

(II),

utilizing combined

gas c h r o m a t o g r a p h i c - m a s s

spectro-

m e t r i c analysis of hexane extracts of these substrates. T h i s r e p o r t c o n s t i t u t e d a d e t a i l e d a c c o u n t of t h e a n a l y t i c a l aspects o f a n e p i d e m i c of i n f a n t deaths associated w i t h the i n g e s t i o n of this c o m p o u n d , w h i c h w a s the a c t i v e i n g r e d i e n t of a m i l d e w p r e v e n t a t i v e , the suspected source of exposure.

A n a l y s i s of the hexane extracts w a s

p e r f o r m e d o n a 1.0 m X 2.5 m m c o i l e d glass gas c h r o m a t o g r a p h i c c o l u m n packed with 3 %

D E G S and 2 %

concentrated phosphoric acid.

Under

these c o n d i t i o n s , analysis c o u l d b e p e r f o r m e d w i t h o u t p r i o r d e r i v a t i z a t i o n of the P C P . A l l s a m p l e spectra gave f o u r i d e n t i c a l m a j o r ions w h e n c o m p a r e d w i t h P C P s t a n d a r d mass s p e c t r u m o b t a i n e d at 20 e V . T h u s , i d e n t i t y of the residues of P C P was c o n f i r m e d o n the basis of the o b s e r v a t i o n of t h e m o l e c u l a r i o n p e a k at m / e 264, a f r a g m e n t i o n of m / e 229, o w i n g t o e l i m i n a t i o n of a C I a t o m , a m a j o r f r a g m e n t r e s u l t i n g f r o m the c o n s e c u t i v e loss of H C 1 a n d C O at m / e 200, a n d finally a c h a r a c t e r i s t i c f r a g m e n t of

0

2

4

6

8

1

0

1

2

1

4

0

2

4

6

8

H>

U

ft

Figure I . Total ion current chromatograms of (A) standard 1-naphthyl chloroacetate (1 fig) and (B) 1-naphthyl chloroacetate isolated from human urine Programmed temperature conditions: two minutes at 165°C, isothermal at 185°C

to 185°C

at


5°C/min,

136

PESTICIDES

IDENTIFICATION


-CHCICO

B

Figure

m/e

2.

Electron

/.89

impact-induced fragmentation chloroacetate

scheme for

1-naphthyl

165 u n d o u b t e d l y r e s u l t i n g f r o m loss of a c h l o r i n e a t o m f r o m the

m / e 200 i o n . D u r i n g the course o f a n a l y t i c a l m e t h o d studies i n v o l v i n g t h e d e v e l o p m e n t of p r o c e d u r e s for the d e t e c t i o n a n d q u a n t i t a t i o n of 1 - n a p h t h o l , a m a j o r m e t a b o l i t e of c a r b a r y l , i n h u m a n a n d a n i m a l u r i n e ( 1 2 ) ,

the

d e r i v a t i z e d r e s i d u e , 1 - n a p h t h y l chloroacetate, w a s c o n f i r m e d b y gas c h r o matography-mass spectrometry. l a t i o n of

T h e a n a l y t i c a l p r o c e d u r e f o r t h e iso-

1 - n a p h t h o l residues i n v o l v e d a c i d h y d r o l y s i s of

1-naphthol

conjugates i n t h e u r i n e , b e n z e n e e x t r a c t i o n , d e r i v a t i z a t i o n w i t h c h l o r o acetic a n h y d r i d e a n d p y r i d i n e , a n d r e m o v a l of i n t e r f e r i n g m a t e r i a l i n urine b y c o l u m n chromatography using silica gel. 1 - N a p h t h y l chloroacetate w a s e l u t e d w i t h 6 0 % b e n z e n e - h e x a n e .

P r o g r a m m e d t e m p e r a t u r e gas

c h r o m a t o g r a p h i c analysis w i t h a c o i l e d glass c o l u m n , 4 ft X 1 / 8 i n c h o.d., p a c k e d w i t h 2 % S E - 3 0 o n 6 0 / 8 0 m e s h G a s C h r o m Q , w a s u s e d to c o n f i r m residues of 1 - n a p h t h o l as the chloroacetate ester i n h u m a n u r i n e of i n d i v i d u a l s o c c u p a t i o n a l l y e x p o s e d to c a r b a r y l ( F i g u r e 1 ) .

D i a g n o s t i c mass

s p e c t r a l peaks o b s e r v e d for 1 - n a p h t h y l chloroacetate i n c l u d e d the m o l e c u l a r i o n of m / e 220, a base p e a k f r a g m e n t w h i c h m a y b e f o r m u l a t e d as a 1 - n a p h t h o l r a d i c a l i o n ( o r a n i s o m e r ) f o u n d at m / e

144 a r i s i n g f r o m

h y d r o g e n r e a r r a n g e m e n t a n d loss of — C H C I C O , a r e l a t i v e l y w e a k p e a k at m / e 127 f o r m e d either b y e l i m i n a t i o n of — O C O C H C l f r o m t h e m o 2

l e c u l a r i o n a n d / o r b y loss of w a t e r f r o m t h e m / e 145 f r a g m e n t , a n d

finally,

other peaks c h a r a c t e r i s t i c of the f r a g m e n t a t i o n of 1 - n a p h t h o l at m / e


115

9.

BIROS


137

Spectrometry

a n d m / e 116 a r i s i n g f r o m e x p u l s i o n of C H O a n d C O , r e s p e c t i v e l y , a n d a p e a k at m / e 89 p r e s u m a b l y r e s u l t i n g f r o m a C H 7

5

+

ion formed b y the

loss of acetylene ( C H ) f r o m t h e m / e 115 f r a g m e n t ( F i g u r e 2 ) . 2

2

Organophosphorus Pesticides I n c o n n e c t i o n w i t h the d e v e l o p m e n t o f a n a n a l y t i c a l m e t h o d f o r t h e d e t e r m i n a t i o n of o r g a n o p h o s p h o r u s

(13)

pesticides i n h u m a n b l o o d

a n d u r i n e , mass s p e c t r a l c o n f i r m a t i o n of a series of m e t h y l a t e d a n d e t h y l a t e d d e r i v a t i v e s of t h e h y d r o l y t i c a n d m e t a b o l i c p r o d u c t s of these i n s e c t i Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

cides w a s r e q u i r e d . T h e u r i n e o f a n i n d i v i d u a l o c c u p a t i o n a l l y

exposed

to p a r a t h i o n w a s e x t r a c t e d w i t h a 1:1 ( v / v ) solvent m i x t u r e of a c e t o n i t r i l e a n d d i e t h y l ether. S i m u l t a n e o u s l y , t h e i n t a c t o r g a n o p h o s p h o r u s i n s e c t i cides w e r e h y d r o l y z e d b y a d d i n g a p o r t i o n o f 5 N h y d r o c h l o r i c a c i d t o

O

2

4

6

8

K>

12

O

2

4

6

8

10

12

14

14

16

Figure 3. Total ion current chromatograms of (A) standard mixture of methylated dialkylphosphateSy phosphorothioates, and phosphorodithioates (2 fig) and (B) human urine extract containing methylated hydrolytic and metabolic products of parathion Programmed temperature conditions: five minutes at 75°C, to 120°C at 5°C/min, isothermal at 120°C


138

PESTICIDES

IDENTIFICATION

the u r i n e . F o l l o w i n g e x t r a c t i o n , the o r g a n i c l a y e r w a s c o n c e n t r a t e d a n d t h e residues i n s o l u t i o n d e r i v a t i z e d b y means of d i a z o m e t h a n e . T h e m i n e extract was t h e n s u b j e c t e d to a c l e a n - u p p r o c e d u r e e m p l o y i n g s i l i c a g e l c o l u m n c h r o m a t o g r a p h y (14),

and benzene ( I ) , 4 0 %

e t h y l acetate i n

b e n z e n e ( I I ) , a n d 8 0 % e t h y l acetate i n b e n z e n e ( I I I ) as e l u t i n g solvents. C o n c e n t r a t i o n of t h e c h r o m a t o g r a p h i c c o l u m n eluents p r o v i d e d the u r i n e extract w h i c h w a s subjected to p r o g r a m m e d t e m p e r a t u r e analyses o n a c o i l e d glass c a p i l l a r y c o l u m n , 110 ft X 0.025 i n c h i . d . , c o a t e d w i t h V e r s a m i d 900 c o n t a i n i n g 5 % I g e p a l C O - 8 8 0 . T h e r e s u l t i n g t o t a l i o n c u r r e n t c h r o m a t o g r a m is s h o w n i n F i g u r e 3. A l s o i l l u s t r a t e d is the t o t a l i o n c u r r e n t Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

trace o b t a i n e d b y analysis of a series of m e t h y l a t e d d i a l k y l

phosphate

standard materials. T h e m e t a b o l i c a n d / o r h y d r o l y t i c p r o d u c t s of p a r a t h i o n e n c o u n t e r e d as residues i n t h e u r i n e i n c l u d e b o t h d i e t h y l p h o s p h o r i c a c i d a n d d i e t h y l p h o s p h o r o t h i o i c a c i d , most p r o b a b l y as t h e i r salts ( p o t a s s i u m or s o d i u m ) . D e r i v a t i z a t i o n of these residues w i t h d i a z o m e t h a n e w o u l d result i n the f o r m a t i o n of three t r i a l k y l phosphate c o m p o u n d s , n a m e l y , 0 , 0 - d i e t h y l O-methyl thionate

phosphate (DEMMTP),

(DEMMPTh).

(DEMMP), and

0,0-diethyl

0,0-diethyl

O-methyl

S-methyl

phosphoro-

phosphorothiolate

E a r l i e r ( 1 5 ) , it h a d b e e n s h o w n b y c o m b i n e d gas c h r o -

m a t o g r a p h y - m a s s s p e c t r o m e t r y a n d other a n a l y t i c a l d a t a that a l a t e r e l u t i n g major p r o d u c t

(ca.

85%)

of the m e t h y l a t i o n of d i e t h y l p h o s -

p h o r o t h i o i c a c i d f o r m e d u n d e r the c o n d i t i o n s of the a n a l y t i c a l m e t h o d was

DEMMPTh,

DEMMTP.

and

the

minor

product

formed

(ca.

15%)

was

A c c o r d i n g l y , a l l three t r i a l k y l phosphates w e r e o b s e r v e d a n d

c o n f i r m e d b y mass s p e c t r o m e t r y i n the analysis of the h u m a n u r i n e extract. Sufficient i n t e r n a l b o n d energy differences are associated w i t h the i s o m e r i c structures D E M M P T h

and D E M M T P

that q u a l i t a t i v e l y a n d

q u a n t i t a t i v e l y d i s s i m i l a r f r a g m e n t a t i o n patterns are o b s e r v e d for

both

isomers as c a n b e seen f r o m t h e mass spectra of these c o m p o u n d s s h o w n i n F i g u r e 4. T h e mass s p e c t r u m of D E M M T P is c h a r a c t e r i z e d b y b o t h p h o s p h o r u s - o x y g e n a n d c a r b o n - o x y g e n b o n d fission as e v i d e n c e d b y the f o r m a t i o n of f r a g m e n t ions at m / e 140 ( M - G > H 0 ) a n d m / e 156 4

(M-C H ). 2

4

S i n g l e a n d d o u b l e h y d r o g e n rearrangements a c c o m p a n y the loss of the e t h y l substituent. S u b s e q u e n t f r a g m e n t a t i o n reactions n o t e d w e r e s i m i l a r b o n d fission i n v o l v i n g e l i m i n a t i o n of the r e m a i n i n g a l k y l substituents as a n e t h y l e n e m o l e c u l e or a n e t h o x y l r a d i c a l . A r e l a t i v e l y intense peak, for e x a m p l e , of m / e 111 [ C H * O P ( : S ) - O H ] , appears to b e f o r m e d b y c o n +

secutive loss of ethylene ( t o f o r m the m / e 156 i o n ) a n d a n e t h o x y l s u b stituent f r o m t h e m o l e c u l a r i o n as e v i d e n c e d b y metastable peaks at m / e 132.26 a n d m / e 78.98 o b s e r v e d for these t w o processes. Significant f r a g m e n t ions w e r e also f o r m e d t h r o u g h loss of b o t h s u l f u r atoms or s u l f h y d r y l


9.

BIROS


139

Spectrometry CH C M

100

t

79 . CH^OPOH

/

N

CH CH 0 {

90

2

/

,SHI HO'

Figure 5. Electron impact-induced fragmentation scheme for O O-diethyl O-methyl phosphorothionate (DEMMTP) y

C¥H 0 j>

T

N /

2

CH CH 0 3

2

-C H 0 2

.»

/

m/t 184 2

3

2

3

3

HO

~]t

HO' m/t 112 -C2H4

CH CH 0-P 3

2

2

m/t 138 CH CH 0^ / / " | P-SCH HO' m/t 156

4

1 3

2

CH CH 0. "It >-0H« CH CH 0'

4

-C H 2

CH CH0-P-SCHj 0-P-SCH ' n / i 140 3

-C H

-it

4

•/•

3

^0H

110

2

x

3

CH0 2

5

0 H0-P-SCH

3

m/t 111

m/t 126

CH CH=0-P=0 M/I 91 3

Figure

6. Electron impact-induced fragmentation scheme for ethyl S-methyl phosphorothiolate (DEMMPTh)

0,0-di-


u

9.

BIROS


compounds process A

141

Spectrometry

h a v e also b e e n o b s e r v e d

to u n d e r g o

this r e a r r a n g e m e n t

(18). somewhat more complex dissociative scheme was observed

DEMMPTh

(Figure 6).

T h e molecular i o n of this c o m p o u n d

poses b y fission of a c a r b o n - o x y g e n b o n d ( m / e

156, M - C H ) , 2

for

decom-

4

phos-

p h o r u s - o x y g e n b o n d ( m / e 140, M - C H 0 ) , a n d the p h o s p h o r u s - s u l f u r 2

b o n d ( m / e 138, M - C H S ) . 2

4

T h e s t r u c t u r e of t h e m / e 156 f r a g m e n t ( F i g -

ure 6) permits a subsequent identical triple decomposition scheme to y i e l d fragments o f m / e 128 ( 1 5 6 - C H ) , m / e 109 ( 1 5 6 - C H S ) , a n d m / e 2

111 ( 1 5 6 - C H 0 ) . 2

5

4

3

A n o t h e r significant r e a c t i o n o b s e r v e d w a s the e l i m i -


n a t i o n o f w a t e r w h i c h , for e x a m p l e , l e d to t h e f o r m a t i o n of a n intense i o n f r a g m e n t at m / e 91 ( 1 0 9 - H O ) , c h a r a c t e r i s t i c o f t h e e l e c t r o n i m p a c t 2

f r a g m e n t a t i o n of esters. T h e d e c o m p o s i t i o n of D E M M P u n d e r e l e c t r o n i m p a c t results i n t h e f o r m a t i o n of significant f r a g m e n t ions of m / e 141, m / e 113, a n d m / e 9 5 , a c c o r d i n g to the d i s s o c i a t i v e s c h e m e i l l u s t r a t e d i n F i g u r e 7.

Organochlorine Pesticides C h l o r i n a t e d h y d r o c a r b o n p e s t i c i d e residues i n h u m a n a d i p o s e tissue a n d l i v e r tissue samples h a v e b e e n i d e n t i f i e d b y mass s p e c t r o m e t r y c o u pled with

gas c h r o m a t o g r a p h y .

A

g e n e r a l , extensive

extraction a n d

c l e a n u p p r o c e d u r e a d a p t e d f r o m e x i s t i n g m e t h o d s w a s u s e d to isolate

Figure

7.

Electron

impact-induced fragmentation scheme for O-methyl phosphate (DEMMP)

O O-diethyl


y

142

PESTICIDES

IDENTIFICATION

a n d p u r i f y the o r g a n o c h l o r i n e residues. T h e tissue samples w e r e e x t r a c t e d w i t h a c e t o n i t r i l e o r p e t r o l e u m ether, f o l l o w e d b y l i q u i d - l i q u i d p a r t i t i o n i n g o f t h e extract b e t w e e n hexane or p e t r o l e u m ether a n d a c e t o n i t r i l e . C o l u m n chromatography and cleanup on Florisil provided t w o individual fractions ( 1 9 ) , w h i c h w e r e t h e n c o n c e n t r a t e d a n d subjected t o f u r t h e r c l e a n u p a n d p e s t i c i d e r e s i d u e f r a c t i o n a t i o n b y means o f s i l i c a g e l c o l u m n c h r o m a t o g r a p h y e m p l o y i n g a p u b l i s h e d p r o c e d u r e (20).

T h e pesticide

residues w e r e separated b y t h e s i l i c a g e l c h r o m a t o g r a p h i c step i n t o t w o fractions w i t h sufficient r e s o l u t i o n o n t h e gas c h r o m a t o g r a p h i c c o l u m n for t h e i d e n t i f i c a t i o n o f seven p e s t i c i d e residues f r o m t h e a d i p o s e tissue Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

s a m p l e a n d six p e s t i c i d e residues f r o m t h e l i v e r tissue sample. i n c l u d e d : /?- a n d y - h e x a c h l o r o c y c l o h e x a n e

(HCH),

These

1,4,5,6,7,8,8-hepta-

chloro-2,3-epoxy-2,3,3a,4,7,7a-hexahydro-4,7-methanoindene

(heptachlor

e p o x i d e ) , 1,2,3,4,10,10 - h e x a c h l o r o - 6,7 - e p o x y - l,4,4a,5,6,7,8,8a - o c t a h y d r o 1,4 - endo,exo - 5,7 - d i m e t h a n o n a p h t h a l e n e p h e n y l ) -1,1-dichloroethylene

(dieldrin),

(p,p'-DDE),

2,2 - b i s (p - c h l o r o -

2,2-bis ( p - c h l o r o p h e n y l ) -1,1-

dichloroethane ( p , p ' - D D D ) , a n d l,l-bis(p-chlorophenyl)-2,2,2-trichloroethane

a n d 1- ( o - c h l o r o p h e n y l ) - 1 - ( p - c h l o r o p h e n y l ) -2,2,2-trichloroethane

( p , p ' - D D T a n d o , p ' - D D T , respectively). T h e concentration of the pesticides i n tissue r a n g e d f r o m 0.073 to 28.7 p p m . G a s c h r o m a t o g r a p h i c separations w e r e p e r f o r m e d o n a 4 ft X 1 / 8 i n c h o.d. c o i l e d glass c o l u m n packed w i t h 3 % O V - 2 1 0 on 80/100 mesh Chromosorb W ( H P ) . F i g u r e 8 illustrates t h e t o t a l i o n c u r r e n t c h r o m a t o g r a m o b t a i n e d b y analysis of t h e a d i p o s e tissue extract r e p r e s e n t i n g 5 grams of tissue. I n s t r u m e n t a l s e n s i t i v i t y w a s sufficient t o i d e n t i f y residues at c o n c e n trations of 0.30 p p m . C o m p u t e r e n h a n c e m e n t t e c h n i q u e s (21) p e r m i t t e d i d e n t i f i c a t i o n of h e p t a c h l o r e p o x i d e residues at a l e v e l of 0.073 p p m .

__,

0

,

2

-

l

4

6

8

i

10 12

14

MINUTES

~6

2

4

6

8

10

12 14 MINUTES

Figure 8. Total ion current chromatograms of extracts containing organochh rine pesticide residues isolated from adipose tissue (A) fraction 1 and (B) fraction 2 Programmed temperature conditions: one minute at 150°C, min, isothermal at 200°C

to 200°C

at 5°CI


9.

BIROS


Spectrometry

143

D e f i n i t i v e c o n f i r m a t i o n of p e s t i c i d e residues w a s o b t a i n e d b y c o m p a r i s o n of p a r e n t a n d f r a g m e n t i o n intensities a n d mass n u m b e r s of e l u t e d p e s t i cides a n d reference pesticides. T a b l e I lists the residues e n c o u n t e r e d a n d t h e mass n u m b e r s a n d intensities of t h e c h a r a c t e r i s t i c fragments e m p l o y e d for i d e n t i f i c a t i o n i n the a d i p o s e tissue s a m p l e . T h e mass s p e c t r a l f r a g m e n t a t i o n patterns for a l l t h e c o m p o u n d s i n c l u d e d i n T a b l e I w i t h t h e e x c e p t i o n of / 2 - H C H h a v e b e e n a d e q u a t e l y d i s c u s s e d b y other i n v e s t i -


gators

(7).

Table I. Gas Chromatographic Peak Identities and Characteristic Mass Spectral Fragments and Intensities of Some Organochlorine Pesticide Residues Isolated From H u m a n Adipose Tissue Peak

No.

Pesticide p,p'-DDE

1

2

3

o,p'-DDT

p,p'-DDT

£-HCH

5

6

H e p t a c h l o r epoxide

Dieldrin

p,p'-DDD

Characteristic Mass Spectral and Intensities

Peaks

m / e 246 ( 1 0 0 % ) , 318 ( 8 3 % ) , 316 ( M o l e c u l a r i o n , 6 6 % ) , 248 ( 5 8 % ) , 320 ( 4 1 % ) , 176 ( 4 1 % ) , 210 ( 1 6 % ) . m / e 2 3 5 ( 1 0 0 % ) , 2 3 7 ( 7 0 % ) , 165 ( 4 0 % ) , 75 ( 2 0 % ) , 199 ( 1 8 % ) , 246 ( 1 8 % ) , 352 (Molecular ion, 3%). m / e 235 ( 1 0 0 % ) , 237 ( 7 2 % ) , 165 ( 4 8 % ) , 75 ( 2 2 % ) , 50 ( 1 8 % ) , 51 ( 9 % ) , 352 (Molecular ion, 2 % ) . m / e 109 ( 1 0 0 % ) , 2 1 9 ( 8 5 % ) , 181 ( 7 8 % ) , 183 ( 8 0 % ) , 111 ( 6 2 % ) , 193 ( 6 2 % ) , 288 ( M o l e c u l a r i o n , 4 % ) . m / e 81 ( 1 0 0 % ) , 353 ( 8 4 % ) , 355 ( 7 6 % ) , 351 ( 4 8 % ) , 357 ( 3 5 % ) , 237 ( 3 3 % ) , 386 ( M o l e c u l a r i o n , 8 % ) . m / e 79 ( 1 0 0 % ) , 108 ( 1 9 % ) , 263 ( 1 8 % ) , 277 ( 1 9 % ) , 279 ( 1 6 % ) , 345 ( 7 % ) , 378 (Molecular ion, 3%). m / e 2 3 5 ( 1 0 0 % ) , 2 3 7 ( 6 6 % ) , 165 ( 5 8 % ) , 75 ( 2 1 % ) , 82 ( 1 6 % ) , 88 ( 1 6 % ) , 318 (Molecular ion, 5%).

S o m e difficulties w e r e e n c o u n t e r e d i n the absolute c o n f i r m a t i o n of c o n f i g u r a t i o n a l a n d s t r u c t u r a l isomers w h e r e mass s p e c t r a l p e a k intensities a n d mass values of c h a r a c t e r i s t i c f r a g m e n t ions w e r e too s i m i l a r u n d e r the c o n d i t i o n s of the analysis. I n those instances, h o w e v e r , i d e n t i f i c a t i o n w a s r e a d i l y m a d e o n the basis of gas c h r o m a t o g r a p h i c r e t e n t i o n t i m e d a t a . T h e e s t i m a t e d l o w e r l i m i t of d e t e c t a b i l i t y of o r g a n o c h l o r i n e p e s t i c i d e residues b y this a n a l y t i c a l scheme a n d i n s t r u m e n t c o n f i g u r a t i o n w a s d e t e r m i n e d to b e a p p r o x i m a t e l y 0.05 to 0.1 p p m .


144

PESTICIDES

IDENTIFICATION

Herbicide Residues C o m b i n e d m e t h o d o l o g y a n d m e t a b o l i s m studies i n v o l v i n g t h e h e r b i cides 2,4-dichlorophenoxyacetic a c i d ( 2 , 4 - D ) a n d 2,4,5-trichlorophenoxya c e t i c a c i d ( 2 , 4 , 5 - T ) f u r n i s h e d a n o p p o r t u n i t y f o r the a p p l i c a t i o n o f the gas c h r o m a t o g r a p h i o - m a s s s p e c t r o m e t r i c t e c h n i q u e t o the c o n f i r m a t i o n of d e r i v a t i z e d i n t a c t residues a n d m e t a b o l i t e s a n d t h e s t r u c t u r a l c h a r a c terization o f u n k n o w n metabolites isolated from rat urine. T h e analytical technique, briefly summarized, involves a c i d hydrolysis of phenolic conjugates, e x t r a c t i o n o f free p h e n o l s a n d acids w i t h b e n z e n e , extract c o n -


centration

and

derivatization with

diazoethane,

and

chromatographic cleanup a n d separation b y silica gel employing benzene-hexane as eluents ( 2 2 ) .

finally,

column

chromatography

a n d b e n z e n e - e t h y l acetate solvent

systems

P r o g r a m m e d t e m p e r a t u r e gas c h r o m a t o g r a p h i c analyses

of t h e u r i n e extracts w i t h a 7 ft X 1 / 8 i n c h o.d. a l u m i n u m c o l u m n p a c k e d w i t h 2 0 % O V - 1 0 1 on 6 0 / 8 0 mesh Gas C h r o m Q p r o v i d e d the total ion c u r r e n t c h r o m a t o g r a m s s h o w n i n F i g u r e 9. T h e mass s p e c t r a l f r a g m e n t a t i o n d a t a w e r e u s e d t o c o n f i r m the presence o f 2 , 4 - D a n d 2,4,5-T as t h e e t h y l esters a n d t r i c h l o r o p h e n o l ( T C P ) as the e t h y l ether i n the s e v e r a l

"12.4.5-TCP

•

2

4

•

t

M

1

2

W

t

t

t

«

2

*

2

2

2

4

Figure 9. Total ion current chromatograms of animal urine extracts containing (A) 2,4-D, (B) 2,4,5-T, and (C) 2,4,5-trichlorophenol and an isomer of trichlorodihydroxybenzene, peak 2 (as ethyl esters and mono- and diethyl ethers, respectively) Programmed temperature conditions: five minutes at 180° C, to 220° C at 5°C/min, isothermal at 220°C


9.

BIROS


Spectrometry

^

^

145

OCRjCOOCijClj

• / • 213

— I -C«2=CH 2

CI

• / , 24S


-COOCt • / • "5

»

C H 0Cl7 6

•/

4

t

162

Figure 10. Electron impact-induced fragmentation scheme for 2,4-dichlorophenoxyacetic acid, ethyl ester analyses p e r f o r m e d .

T h e diagnostic fragmentation pattern for 2,4-D

e t h y l ester i n c l u d e d peaks at m / e 248, m o l e c u l a r i o n ; m / e 213, e l i m i n a t i o n of a C I a t o m ; m / e 185, c o n s e c u t i v e e l i m i n a t i o n o f a C I a t o m a n d e t h y l e n e ( C H ) f r o m t h e p a r e n t i o n species; m / e 175, loss o f t h e c a r b o e t h o x y l 2

4

f u n c t i o n f r o m t h e m o l e c u l a r i o n ; a n d finally, a p e a k a t m / e 162 p r e s u m ably arising from a dichlorophenol or dichlorocyclohexadienone-type

rad-

i c a l i o n ( F i g u r e 1 0 ) . A n analogous f r a g m e n t a t i o n s c h e m e w a s o b s e r v e d for t h e e t h y l ester o f 2,4,5-T: m / e 282, m o l e c u l a r i o n ; m / e 2 4 7 , loss o f c h l o r i n e a t o m ; m / e 219, consecutive e l i m i n a t i o n o f a c h l o r i n e a t o m a n d a m o l e c u l e o f e t h y l e n e f r o m t h e p a r e n t i o n ; m / e 209, d e c o m p o s i t i o n o f the p a r e n t i o n w i t h e l i m i n a t i o n o f t h e c a r b o e t h o x y l g r o u p ; a n d m/e

196, c o r r e s p o n d i n g

finally,

to a trichlorophenol or trichlorocyclohexa-

d i e n o n e - t y p e r a d i c a l i o n species. I n a d d i t i o n to t h e i n t a c t residues o f 2,4,5-T excreted i n t h e r a t u r i n e , mass s p e c t r a l e v i d e n c e w a s o b t a i n e d f o r t h e presence o f t h e m e t a b o l i t e T C P a n d a t r i c h l o r o d i h y d r o x y b e n z e n e isomer, w h i c h w e r e o b s e r v e d as t h e i r m o n o - a n d d i e t h y l ether d e r i v a t i v e s , r e s p e c t i v e l y . T h u s , T C P e t h y l ether d i s p l a y e d a m o l e c u l a r i o n at m / e 224 a n d c h a r a c t e r i s t i c fragments at m / e 196 a n d m / e 160 o w i n g to consecutive e l i m i n a t i o n o f e t h y l e n e a n d H C 1 f r o m t h e p a r e n t i o n . T h e f r a g m e n t a t i o n p a t t e r n o f 2,4,5-tric h l o r o p e n o l w a s o b s e r v e d t o e x h i b i t s i m i l a r b e h a v i o r b e l o w m / e 196.


146

•

PESTICIDES

J

4

6

8

10 U

M

16 O

minutes

20 22 24 2^

~0

2

4

6

8

IDENTIFICATION

10121416182022242628

minutes


Figure 11. Total ion current chromatograms of (A) standard Aroclor 1260 mixture of polychlorinated biphenyls. Programmed temperature analysis: two minutes at 200°C, to 230°C/min, isothermal at 230°C. (B) Human adipose tissue extract. Programmed temperature analysis: two minutes at 190°C, to 230°C at 5°C/min, isothermal at 230°C.

T h e mass s p e c t r a l e v i d e n c e f o r t h e t r i c h l o r o d i h y d r o x y b e n z e r r e d i e t h y l ether i n c l u d e d a m o l e c u l a r i o n o f m / e 286 a n d significant f r a g m e n t ions at m / e 240, m / e 212, a n d m / e 176 a r i s i n g f r o m the c o n s e c u t i v e e l i m i n a t i o n o f t w o m o l e c u l e s o f e t h y l e n e a n d one m o l e c u l e o f H C 1 .

Polychlorinated Biphenyls C o n f i r m a t o r y e v i d e n c e f o r the presence o f p o l y c h l o r i n a t e d b i p h e n y l s ( P C B ) i n fish, seabirds, c o n i f e r needles, a n d h u m a n d e p o t f a t has b e e n o b t a i n e d b y gas c h r o m a t o g r a p h y - m a s s s p e c t r o m e t r y (23, 24). T h e mass s p e c t r a l b e h a v i o r o f standards o f P C B m a t e r i a l s ( A r o c l o r 1254 a n d 1260) a n d P C B c o m p o u n d s i s o l a t e d f r o m h u m a n a d i p o s e tissue u t i l i z i n g s t a n d ard analytical methodology

(19) h a s b e e n r e p o r t e d r e c e n t l y ( 2 5 ) .

illustration of the total i o n current chromatogram

An

obtained b y pro-

g r a m m e d t e m p e r a t u r e analysis o f a m i x t u r e of s t a n d a r d P C B c o m p o u n d s ( A r o c l o r 1260) o n a 100 ft X 0.02 i n c h i . d . stainless steel c a p i l l a r y c o l u m n c o a t e d w i t h O V - 1 s i l i c o n e o i l is d e p i c t e d i n F i g u r e 11. M o s t c o m p o n e n t s of the A r o c l o r s t a n d a r d p r o d u c e d mass spectra w i t h m o l e c u l a r i o n groups of h i g h i n t e n s i t y . I n a d d i t i o n , t h e c h a r a c t e r i s t i c isotopic d i s t r i b u t i o n patterns c o r r e s p o n d i n g t o t h e n u m b e r o f c h l o r i n e atoms i n the p a r e n t i o n a n d c h l o r i n e - c o n t a i n i n g f r a g m e n t ions w e r e o b s e r v e d .

Other noteworthy

features o f t h e mass spectra w e r e t h e r e l a t i v e l y intense i o n fragments r e s u l t i n g f r o m consecutive loss o f c h l o r i n e atoms f r o m the p a r e n t i o n a n d the presence o f intense d o u b l y c h a r g e d fragments w i t h i n the mass spectra of most o f the P C B c o m p o u n d s .

T h u s , i t was possible to establish t h e

m o l e c u l a r w e i g h t a n d n u m b e r o f c h l o r i n e atoms associated w i t h major numbered component

i n t h e c h r o m a t o g r a m o f F i g u r e 11.

each The

s t a n d a r d A r o c l o r 1260, o n this basis, w a s s h o w n t o consist o f at least t w o p e n t a c h l o r o b i p h e n y l s , f o u r h e x a c h l o r o b i p h e n y l s , six h e p t a c h l o r o b i phenyls, t w o octachlorobiphenyls, t w o nonachlorobiphenyls, a n d decachlorobiphenyl.


9.

BIROS


147

Spectrometry

A l t h o u g h i t w a s a p p a r e n t that several of the m a j o r peaks i n the c h r o m a t o g r a m of the s t a n d a r d P C B m a t e r i a l w e r e i n fact c o m p o s e d of m o r e t h a n a single c o m p o n e n t , n o f u r t h e r a t t e m p t w a s m a d e to elaborate o n the i d e n t i t y or n o n i d e n t i t y of the i n d i v i d u a l c o m p o n e n t s of e a c h of these peaks. S t r u c t u r a l studies of t h e P C B c o m p o n e n t s m u s t necessarily i n v o l v e the a p p l i c a t i o n of c o m p l e x s e p a r a t i o n p r o c e d u r e s c o u p l e d w i t h d e t a i l e d s p e c t r o m e t r i c studies of e a c h i s o l a t e d c o m p o n e n t .

F i g u r e 11 also i n c l u d e s

the t o t a l i o n c u r r e n t c h r o m a t o g r a m of a h u m a n a d i p o s e tissue extract w h i c h w a s s h o w n to c o n t a i n traces of P C B c o m p o u n d s ( p e a k s a t h r o u g h Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

/) w h o s e mass spectra w e r e consistent w i t h those f o u n d for A r o c l o r 1260, n a m e l y , peaks 6 a n d 9 t h r o u g h 17. PCB identified

materials i n utilizing

field-collected

combined

gas

b a l d eagle specimens chromatography-mass

have

spectrometry

techniques i n conjunction w i t h thin-layer chromatographic of s t a n d a r d m a t e r i a l s as w e l l as tissue extracts ( 2 6 ) .

been

separations

A t o t a l of 19 P C B

c o m p o u n d s w e r e d e t e c t e d i n the field samples. A n a l y s e s w e r e p e r f o r m e d e m p l o y i n g a s p i r a l glass gas c h r o m a t o g r a p h i c c o l u m n , 9 ft X

0.25 i n c h

o.d., p a c k e d w i t h 1 % S E - 3 0 o n 1 0 0 / 1 2 0 m e s h G a s C h r o m Q . M a s s spectra of the m a j o r c o m p o n e n t s of A r o c l o r 1254 P C B s t a n d a r d w e r e p r e s e n t e d i n this r e p o r t as w e l l as r e l a t i v e r e t e n t i o n d a t a for the i n d i v i d u a l c o m ponents.

Discussion O f a l l the systems w h i c h h a v e b e e n u t i l i z e d for the analysis of p e s t i c i d e residues, c o m b i n e d gas c h r o m a t o g r a p h y - m a s s s p e c t r o m e t r y

affords

a p a r t i c u l a r l y u s e f u l a p p r o a c h because p o s i t i v e i d e n t i f i c a t i o n of the c o m ponents o f a m i x t u r e c a n b e m a d e w i t h o u t p r i o r s e p a r a t i o n at sensitivities c o m p a t i b l e w i t h the l i m i t e d q u a n t i t i e s of residues g e n e r a l l y a v a i l a b l e . F r o m the results of recent studies i n v o l v i n g the a p p l i c a t i o n of this t e c h n i q u e , it has b e e n d e m o n s t r a t e d that a v a i l a b l e r e s i d u e a n a l y t i c a l m e t h o d s p r o v i d e efficient i s o l a t i o n a n d a d e q u a t e c l e a n u p of extracts of h u m a n , a n i m a l , a n d e n v i r o n m e n t a l m e d i a i n most cases to p e r m i t gas c h r o m a t o graphic—mass s p e c t r o m e t r i c analyses w i t h m a x i m u m confidence.

Addi-

t i o n a l l y , i t has b e e n s h o w n t h a t this c o m b i n e d t e c h n i q u e w i l l c o n v e n i e n t l y p r o v i d e d e f i n i t i v e a n d c o n c l u s i v e c o n f i r m a t i o n of r e s i d u e i d e n t i t y as w e l l as c h a r a c t e r i z a t i o n of residues a n d t h e i r metabolites of u n k n o w n s t r u c t u r e . I n a d d i t i o n to the a p p l i c a t i o n s r e p o r t e d here, gas c h r o m a t o g r a p h y mass s p e c t r o m e t r y has b e e n e m p l o y e d i n the c h a r a c t e r i z a t i o n of p h o t o c h e m i c a l d e g r a d a t i o n p r o d u c t s of p , p ' - D D T a n d p , p ' - D D E ( 2 7 ) , s y n t h e t i c i n t e r m e d i a t e s i n p , p ' - D D T m e t a b o l i s m studies ( 2 8 ) , a n d t r a n s f o r m a t i o n p r o d u c t s of h e r b i c i d a l c h l o r o a n i l i n e s i n soil

(29).


148

PESTICIDES

IDENTIFICATION

F r o m a t e c h n i q u e s t a n d p o i n t , a d d i t i o n a l studies s h o u l d b e m a d e o n i m p r o v e m e n t s of i n s t r u m e n t a l s e n s i t i v i t y t o a l l o w r o u t i n e c o n f i r m a t i o n of p e s t i c i d e residues at the n a n o g r a m a n d p o s s i b l y t h e p i c o g r a m l e v e l . S o m e gains h a v e a l r e a d y b e e n r e p o r t e d i n this a r e a , u t i l i z i n g e n h a n c e m e n t t e c h n i q u e s (21), m e t h o d s (4).

p u l s e h e i g h t analysis (30, 31),

computer a n d other

Studies documenting c o m p o u n d y i e l d a n d enrichment fac-

tors f o r the s e v e r a l classes of o r g a n i c pesticides a n d v a r i o u s gas c h r o m a tographic—mass

spectrometric

i n t e r f a c i a l systems

under

a variety

of

o p e r a t i n g c o n d i t i o n s are r e q u i r e d . T e c h n o l o g i c a l studies s y s t e m a t i c a l l y d e s i g n e d to e x p l o i t the advantages of c a p i l l a r y a n d other gas c h r o m a Downloaded by FUDAN UNIV on January 4, 2017 | http://pubs.acs.org Publication Date: June 1, 1971 | doi: 10.1021/ba-1971-0104.ch009

t o g r a p h i c c o l u m n s for p e s t i c i d e r e s i d u e analysis are also n e e d e d .

Finally,

d e v e l o p m e n t of a c o m p l e t e r e s i d u e analysis system e m p l o y i n g e i t h e r a n e l e c t r o n c a p t u r e or flame i o n i z a t i o n detector a n d s p l i t t e r a r r a n g e m e n t i n t a n d e m w i t h the m o l e c u l a r separator p e r m i t t i n g s i m u l t a n e o u s mass spect r o m e t r i c i d e n t i f i c a t i o n a n d d e t e c t i o n a n d q u a n t i t a t i o n of p e s t i c i d e r e s i dues f r o m a s i n g l e gas c h r o m a t o g r a p h i c i n j e c t i o n w o u l d offer

obvious

advantages. T h e r e p o r t e d a p p l i c a t i o n s of c o m b i n e d

gas

chromatography-mass

s p e c t r o m e t r y to the analysis of p e s t i c i d e residues h a v e b e e n i n c r e a s i n g i n f r e q u e n c y i n recent years. W i t h d e v e l o p m e n t s i n i n s t r u m e n t t e c h n o l o g y a n d a v a i l a b i l i t y of i n s t r u m e n t a t i o n , i t c a n b e e x p e c t e d that the t e c h n i q u e w i l l b e r e l i e d u p o n to a n e v e n greater extent i n t h e f u t u r e .

Literature Cited (1) (2) (3) (4) (5) (6) (7) (8) (9)

Biemann, K . , "Mass Spectrometry: Organic C h e m i c a l Applications," 1st ed., p. 46 ff., M c G r a w - H i l l , N e w York, 1962. Budzikiewicz, H., Djerassi, C . , W i l l i a m s , D . H., "Mass Spectrometry of Organic Compounds," p. 6 ff., H o l d e n - D a y , San Francisco, 1967. Gunther, F. A . , 'Instrumentation i n Pesticide Residue Determinations," Advan. Pest Control Res. (1962) 5, 191-319. Biros, F . J., "Recent Applications of Mass Spectrometry and C o m b i n e d Gas Chromatography-Mass Spectrometry to Pesticide Residue A n a l y sis," Residue Rev. (1971) i n press. Kantner, T . R., M u m m a , R. O., "Application of Mass Spectroscopy to Pesticide Residue Analysis," Residue Rev. (1966) 16, 138-51. Westlake, W. E., Gunther, F. A., "Advances i n Gas Chromatographic D e tectors Illustrated from Applications to Pesticide Residue Evaluations," Residue Rev. (1967) 18, 175-217. Sphon, J . A., D a m i c o , J . N., " T h e Mass Spectra of Some Chlorinated Aromatic Pesticidal Compounds," Org. Mass Spectr. (1970) 3, 5 1 - 6 2 and references cited therein. Robinson, J., Richardson, A., Elgar, K . E., " C h e m i c a l Identity i n U l t r a microanalysis," 152nd National Meeting, ACS, New York, September 1966, paper # A - 0 7 5 . Schechter, M. S., " T h e N e e d for Confirmation," Pestic. Monit. J. (1968) 2, 1.



9.

BIROS


Spectrometry

149

(10) Biros, F . J., Walker, A . C., "Pesticide Residue Analysis in Human Tissue by Combined Gas Chromatography-Mass Spectrometry," J. Agr. Food Chem. (1970) 18, 425-9. (11) Barthel, W . F . , Curley, A., Thrasher, C. L., Sedlak, V . A., "Determination of Pentachlorophenol in Blood, Urine, Tissue, and Clothing," J. Assoc. Offic. Anal. Chemists (1969) 52, 294-8. (12) Shafik, M . T., Sullivan, H . , Enos, H . F . , " A Method for the Determination of 1-Naphthol in Urine," Bull. Environ. Contam. Toxicol. (1971) 6, 34-9. (13) Shafik, M . T . , Enos, H . F . , "Determination of Metabolic and Hydrolytic Products of Organophosphorus Pesticide Chemicals in Human Blood and Urine," J. Agr. Food Chem. (1969) 17, 1186-9. (14) Shafik, M . T . , Bradway, D., " A Cleanup Procedure for the Determination of Low Levels of Alkyl Phosphates, Thiophosphates, and Dithiophosphates in Rat Urine," Joint ACS-Chem. Inst. Canada Intern. Meeting, Toronto, May 1970, paper #PEST 054. (15) Shafik, M . T . , Bradway, D . , Biros, F . J., Enos, H . F . , "Characterization of Alkylation Products of Diethyl Phosphorothioate," J. Agr. Food Chem. (1970) 18, 1174-5. (16) Cooks, R. G., Gerrard, A . F . , "Electron Impact Induced Rearrangements in Compounds Having the P=S Bond," J. Chem. Soc. B (1968) 132733. (17) Jorg, J. R., Houriet, R., Spiteller, G., "Massenspektren von Pflanzenschutzmitteln," Monatsh. Chem. (1966) 97, 1064-87. (18) Biros, F . J., Ross, R. T., "Fragmentation Processes in the Mass Spectra of Trialkylphosphates, Phosphorothionates, Phosphorothiolates, and Phosphorodithioates," 18th Conf. Mass Spectr. Allied Topics, San Francisco, June 1970, paper #G3. (19) Food and Drug Administration, "Pesticide Analytical Manual," Vol. I, Sec. 211, General Methods for Fatty Foods, U.S. Dept. of Health, Education, and Welfare, Washington, D . C., revised July 1969. (20) Kadoum, A. M . , "Application of the Rapid Micromethod of Sample Cleanup for Gas Chromatographic Analysis of Common Organic Pesticides in Ground Water, Soil, Plant, and Animal Extracts," Bull. Environ. Contam. Toxicol. (1968) 3, 65-70. (21) Biros, F . J., "Enhancement of Mass Spectral Data by Means of a Time Averaging Computer," Anal. Chem. (1970) 42, 537-40. (22) Shafik, M . T., Sullivan, H . , " A Method for the Determination of Low Level Exposure to 2,4-D and 2,4,5-T," 7th Inter-Am. Conf. Toxicol. Occupational Med., Miami, August 1970; J. Environ. Anal. Chem. (1971) in press. (23) Koeman, J. H . , ten Noever de Brauw, M . C., de Vos, R. H . , "Chlorinated Biphenyls in Fish, Mussels, and Birds from the River Rhine and the Netherlands Coastal Area," Nature (1969) 221, 1126-8. (24) Widmark, G., "Possible Interference by Chlorinated Biphenyls," J. Assoc. Offic. Anal. Chemists (1967) 50, 1069. (25) Biros, F . J., Walker, A . C., Medbery, A., "Polychlorinated Biphenyls in Human Adipose Tissue," Bull. Environ. Contam. Toxicol. (1970) 5, 317-23. (26) Bagley, G. E., Reichel, W . L . , Cromartie, E., "Identification of Polychlorinated Biphenyls in Two Bald Eagles by Combined Gas Liquid Chromatography-Mass Spectrometry," J. Assoc. Offic. Anal. Chemists (1970) 53, 251-61. (27) Plimmer, J. R., Klingebiel, U . I., Hummer, B. E., "Photo-oxidation of D D T and DDE," Science (1970) 167, 67-9.


150 (28) (29) (30) (31)

PESTICIDES

IDENTIFICATION

M c K i n n e y , J. D., Boozer, E . L., Hopkins, H. P . , Suggs, J . E., "Synthesis and Reactions of a Proposed DDT Metabolite, 2,2-bis(p-Chlorophenyl) Acetaldehyde," Experientia (1969) 25, 897-8. Kearney, P. C., Plimmer, J. R., Guardia, F . B . , " M i x e d Chloroazobenzene Formation i n S o i l , " J. Agr. Food Chem. (1969) 17, 1418-9. Bergstedt, L., W i d m a r k , G . , "Repetitive Scanning i n Gas Chromatograp h y - M a s s Spectrometry," Chromatographia (1970) 3, 5 9 - 6 3 . W i d m a r k , G., Bergstedt, L., Laveskog, A., "Mass Spectrometry of Organochlorine Compounds and Attempts to Reach Positive Identification," I.U.P.A.C. Conference, Appl. Chem. Div., Sittingbourne, Kent, U.K., October 1968. September 3,

1970.


RECEIVED


Pesticides Identification at the Residue Level - ACS Publications

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