Acidic pesticdes

using QuEChERS
method
Acidic pesticides
pH-issue: Ionization of pesticides at low or high pH-values
1 Acids: HX 1 H+ + X-

1 Bases: B + H+ 1 BH+
3 Ionic form prefers to stay in the water phase

pH-range of agricultural
samples: ~2.5 – 7
 %
Recovery
Imazapyr
Clopyralid
Picloram
Benazolin
4-CPA
Imazethapyr
Dicamba
MCPA
2,4-D
Acidic pesticides –

Imazaquin
Naphthylacetic acid
Fluoxypyr
2,4,5-T
Mecoprop
Triclopyr
max. pH 5.5
influence of pH on recovery

2,4,5-TP
123456457895
837 123456 789


2,4-DP

Propyzamid

Ioxynil

Bentazon

Bromoxynil

lower pKa 4 general trend 5 higher pKa Fluazifop

Bromacil

2,4-DB

MCPB
pH
6
pH
5
pH
4
pH
0
20
40
60
80

3
100
120
Acidic pesticides – cleanup issue

Example: Citrate-Buffered
Extract of red currant 12
Sample pH 5.1 9
pH=8.3 pH
mg/mL pH=7.8
8
10
7Losses of acidic 7

components after 8 6

cleanup 6
5

4
pH=3.5
4 3

7Acidic compounds 2,4 2

2
interact with PSA 1,0
0,5 1

0 0
Raw Extract PSA 25 PSA 50
mg/mL mg/mL
mg co-extractives/mL Extract
pH
QuEChERS has to be modified,
to include acidic pesticides

7 Avoid special cleanup with PSA

7 Special conditions during chromatography
 Acidic pesticides – pesticide list
Compound pKa-value Chemical Class acidic structure
2,4,5-T 2,85
2,4,5-TP 2,84
2,4-D 2,73
2,4-DB 4,8
2,4-DP 3,67
4-CPA 3,56
Fenoxaprop-P 3,2 phenoxyalcanoic acid
Fluazifop 3,2
Fluazifop-P-Butyl 3,1
Haloxyfop 2,9
MCPA 3,07
MCPB 4,84
MCPP 3,78
Fluroxypyr 2,94
pyridine herbicide
Triclopyr 3,97
Naphthoxyacetic acid, 2- naphthoxyacetic herbicide
Bromoxynil 3,86
phenolic herbicide
Ioxynil 3,96
Clopyralid 2 pyridine herbicide
Dicamba 1,97 benzoic acid
Imazapyr 1,9
Imazaquin 3,8 imidazolinone herbicide
Imazethapyr 2,1
Quinmerac 4,32 quinolinecarboxylic acid
Picloram 2,3 pyridine herbicide
Bentazone 3,3 unclassified
Fomesafen 2,83 amide herbicide
Imazosulfuron 4 pyrimidinylsulfonylurea herbicides
Thifensulfuron-Methyl 4
Metsulfuron-Methyl 3,3 triazinylsulfonylurea herbicides
Metosulam 4,8 sulfonanilide herbicide
Cycloxydim 4,17 cyclohexene oxime herbicide special case
Fludioxonil no acid
Acidic pesticides – pesticide list

Cycloxydim
7 Three mesomeric structures
7 Difficult chromatogramic behaviour
 Acidic pesticides - findings
N°of samples with positive findings
2004 - 2007

CITRUS & POME

ROOT & TUBER

ALL COMMOD.
LEAFY VEGET.

Annex I 91/414
LEGUMES
CEREALS
FRUITING
Pesticide

VEGET.
FRUIT
Fludioxonil 45 297 651 7 10 3283 Pending
Fluazifop
2,4-D
1
98
125
17
47
7 1
9 85
1
421
314
Out
In Relatively often found:
Haloxyfop 1 44 5 2 6 146 Out
Bentazone 72 1 94 In
MCPA 6 1 81 In
Clopyralid 30 1 49 In
Dicamba 8 4 31 Pending 7 Fluazifop
2,4-DP 14 2 25 In
MCPP
4-CPA
2 2
18
24
18
Pending
Pending
7 2,4-D
Naphthoxyacetic acid, 2- 1 16 18 pending
Bromoxynil 10 16 In
Triclopyr 2 9
Imazaquin 7 Pending
2,4,5-T 1 6 In
Quinmerac 1 3
Fluroxypyr 1 2 In
Ioxynil 2 2 In
2,4-DB 1 In
Cycloxydim 1 Pending
Metosulam 1 1 Pending
2,4,5-TP In
Fenoxaprop-P Pending
Formesafen
Imazapyr Out
Imazethapyr Out
Imazosulfuron In
MCPB Out
Metsulfuron-Methyl
Picloram Out
Thifensulfuron-Methyl In
Interlaboratory Method Validation Study
Modified QuEChERS (without PSA-cleanup)

August 2006:
7 19 Labs expressed interest from 9 Countries
(Germany, Latvia, Spain, Czech Republic, Switzerland,
Finland, Austria, The Netherlands and Great Britain)
7 4 Commodities
(Raisins, Cucumber, Lemon, Wheat Flour)
7 32 Pesticides (LC-MS/MS, ESI pos and neg) with
2 Levels (0.1 and 0.01 mg/kg)
 mean recovery in %

0
20
40
60
80
100
120
140
2,
4,
5-
2, T
4,
5-
TP
2,
4-
D
2,
4-
D
B
2,
4-
D
P
4-
C
Be PA
nt
az
Br on
om e
Spiking Level 0.1 mg/kg

ox
C yn
il
lo
py
C ra
yc l id
lo
xy
di
D m
Fe ica
ILMV 2006 Acids –

no m
xa ba
pr
op
Fl -P
ua Flu
zi az
fo i
p- fop
P-
Bu
Fl ty
ud l
io
xo
Spiking Level 0.01 mg/kg

Fl ni
ur l
ox
Fo y py
m r
es
af
H en
Results (Example Cucumber)

al
ox
yf
Im o p
az
ap
Im yr
az
Im aqu
az in
Im eth
az a py
os r
ul
fu
ro
n
Io
xy
ni
l
M
C
PA
M
C
PB
M
M M C
N PP
ap
et
su eto
ht lfu su
ho ro la
xy n m
ac -Me
et th
ic y
ac l
id
,2
Pi -
Th cl
or
ife Q
ns ui am
ul nm
fu er
ro ac
n-
M
et
hy
Tr l
ic
lo
py
r
 QuEChERS – alkaline hydrolysis

Example 2,4-D

7 Selective systemic herbicide

7 Control of broad leaved weed
7 Plant growth regulator used to prevent premature fruit drop
7 Formulations include free acid, salts, esters
7 May form conjugates
7 Method designed to convert all possible residues to free acid

Acids are often covalently bound to matrix components and

thus their concentration underestimated!
QuEChERS – schematic description with AH

Citrate-Buffered QuEChERS

Weigh 10 g of Frozen Sample

alkaline hydrolysis:
Shake
Add NaOH and store for
Add 10 mL Acetonitrile 30 min at RT,
then neutralize
Add ISTD-Solution
Shake
optionally:
Add 4 g MgSO4 / 1 g NaCl / Citrate Buffer Add NaOH to adjust pH
(pH 5-5.5)
Shake and Centrifuge optionally:
Freeze-out of extracted
Analysis of acidic pesticides by LC-MS/MS fat over night
 8888883 
!8"8#8
Alkaline Hydrolysis for the release of phenoxy-acid pesticides
wheat sample " 8 "
1400 At 70 °C At Room Temperature
1200
Relativ concentration

pH 12 pH 10 pH 10 pH 9
1000 30 min 60 min 30 min 30 min

800 715
606 619
600
400 336
275
200 135 100
0

in
in
in

in

in

in

.
ol
m
m
m

dr
30

0
30

30

60

30

hy
3
/

9/
12
/

/
12

10

10

10

o
w/
pH
pH
pH

pH

pH

pH

S
ER
Ch
uE
Q
Alkaline Hydrolysis –
Level of MCPA in berries

(%$
./ 5#/
($$
)8)8842* 8+,-

'%$

'$$

&%$

&$$

%$

$
88  #88 
Alkaline Hydrolysis –
Level of 2,4-D in different citrus samples

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88  #88 

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