�� !" /�� (XCT)�� !"#�� !"#$%&'(

��

�� !

��

Bernhard Wüst, Christian Sauber

Agilent Technologies GmbH,

Hewlett-Packardstr. 8, 76337

Waldbronn

Germany

Hans (J.) A. van Rhijn

RIKILT -Institute of Food Safety

Bornsesteeg 45,6700 AE Wageningen

the Netherlands

��

�� /�� !"#$�� !/��/�� !"#$%&

�� !"#$%&'()*+,-,./01�� !"#$

�� !"#$%& 0.125 � 0.5 g/Kg�� !"�� !

�� 1 g/kg�� !"#$�%&'

��

�� !"#$%&'()*+,-./0����

�� ! �1995���� !"#$�� !"#$

�� !"#$%�� !"�� !"��#$%&

�� �� !"#$%&'()*+,-�� !"

�� !"#$%&�� !"#$%&'()��

� /�� /�� !"#$%&'()*+ !,��

�� !"#$%&'()*+,-.'/0/12

���� !"#$%&'()*+,- 1��

��

�� /�� /�� !"#$%&'( 1100 XCT��

�� !"#$%�� !"#$%&'()*+��

�� !"#$%&'(MRM)��� NBA-AMOZ �

NBA-AOZ�� !"#$%&'(

NBA-SEMSEM

NBA-AHDAHD

NBA-AMOZ

NBA-AOZ

AMOZ

AOZ

ON+

O_

ON

N

N

OO

O

O N N

N

N

O

OO

O

O

O

N

HN

NOO

N

O

OH2N

ON+

O_

O N

O

ON

N

N

OO

O

H2N

ON

N

NO

O

O

N

NO

NH2

HN

OO

ON+

NO

O_

O

NH2

HN

H2N

O

NHON+

O_

ON N

N

O

O O

NH

H2NO

NH2

HN

�� !

� 1�� !"��� !"# , � 5 mL � 0.2 M �

�� !"#$%�� !50�� !"#$%&'

�(100 mM�� !"#"$%&)��370 �� �

�� !�� !"#$%&'()*+,-./0

�� !�RIKILT�Netherlands���� !"#�

�� !�� !"#$%&'($)*+,-.��

�� !""#$%&'(�� !"#$%&�2�

�� !"#$%�� !"#$%&' ()��

� 1. �� !"#$%&'()*+,'*-./

��500��0.3 M�� !"#$%&�� 2 M�

�� !"#$ pH�� ! 7��� 4 mL�� �

���� !"�� !"#$%&'()*�� !

�� !"#$�� !"#"$%&'()*��

�� !"�� ��500-µL��� �50 mL��

80 mL�� 0.5 mL�� !"�� !"# $% .

�� !"#$%&'(�� !"#$!%�� !

�� !"#$%&'()*+,-)./012�

�� !"#$�� !"

2

�� !

�� !

�� !

�� !

�� �� �� !"#$%"

�� !

�� (Biosolve, 13683502)

���32% (Merck, 100319)

2-�� !"(2-NBA) (Sigma, N6001)

12�� !"#$ (Merck, 106578)

�� ! (Merck, 106498)

��� (Biosolve, 05402602)

�� (Merck, 10063)

�� (Biosolve, 01203502)

�� !�99.5% (Aldrich, 15.193-9)

�� /�� /�� !"�� !"#(HPLC): Agilent 1100

�� �! : 0.4 mL/min

�� : Zorbax XDB-C8, 2.1 mm x 50 mm, 3.5 m

�� : A: H2O + 0.1% ��

B: �� + 0.1% ����: 0-14 min; 10% A -45% A; 14-16 min; 45% A -90% A

�� ! : 500 l �� !" 50 l

1100 LC/MSD XCT �� !"�� ! : ESI �� !"

�� !" : 45 psi

�� !" : 12 L/min

�� !" : 350 �

�� !(Skimmer): 20 V

�� !"#$: 55 V

�� !"( Trap Drive): 55

ICC: on

�� !"#$(MRM)

4 �� !": 0-2.2 min, 2.2-7.4 min, 7.4-10.6 min, 10.6-13.9 min

�� 1: �� !": �� !" , �� !"

�� 2: m/z 335 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.16

m/z 340 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.16

�� 3: m/z 209 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.28

m/z 249 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.25

�� 4: m/z 236 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.25

m/z 240 �MS/MS, Isolation width: 2.0, Cut-off: 140, Amplitude: 1.25

�� !

NBA-AMOZ: EIC of 261 + 291 (m/z 335 � MS/MS), �� !: 4.5 min

NBA-dAMOZ: EIC of 266 + 296 (m/z 340 � MS/MS), �� !: 4.5 min

NBA-SEM: EIC of 166 + 192 (m/z 209 � MS/MS), �� !: 9.9 min

NBA-AHD: EIC of 134 (m/z 249 � MS/MS), �� !: 10.0 min

NBA-AOZ: EIC of 134 (m/z 236 � MS/MS), �� !: 10.8 min

NBA-dAOZ: EIC of 134 (m/z 240 � MS/MS), �� !: 10.8 min

Max Acc time: 150 ms

Smart target: 100000

Scan: 100-350

3

NBA-AMOZ2A 2B

123

x104

0

NBA-SEM

x104

0.0

0.4

0.8

1.2

2 4 6 8 10 12 Time [min]

NBA-d5AMOZ

NBA-AMOZ

NBA-d5AMOZx105

0.0

0.4

0.8

NBA-AHD

0

2000

4000

6000

NBA-AOZ

x104

0.0

1.0

2.0

NBA-d4AOZ

NBA-SEM

NBA-AHD

NBA-AOZ

NBA-d4AOZ

x104

0.0

1.0

2.0

122.9

133.8

148.8158.9

166.7176.8

204.8 222.8 239.9

264.9

295.9

192.9 261.9

290.9

317.8 333.9

135.8148.7

165.7

190.8 207.8217.8 228.7

120.8

133.8

138.7144.8

162.7

170.7188.7

200.7212.8

230.7248.8

119.9

133.8

148.8

164.7

171.8190.7

206.7

217.7 235.8

0246

x104

x104

x104

x104

IntensityIntensity

0

1

2

3

02000400060008000

02000400060008000

0.0

1.0

2.0

0.0

1.0

2.0

3.0

100 125 150 175 200 225 250 275 300 325 m/z

�� !"

�� !"#$%&'()*+,-./01232

�� !�� !"#$%& '()*+,-./

�� !"�� ! "#$%&�� !"#$%&

�� !"#$(LOD)�

�� !"#$�� !"#$%&'()*+,��

�� !"�� !"#$%& !'�� !"#/

�� !"�� !"#$%&'()*+�� !"

�� !"#$%&'("#)*+�� !"#$

�� !"#�� 2�

� 2. �� !"#$%&'()*+ (2A) �� !"#$%&

4

0

1000

2000

3000

4000

2.5 3.0 3.5 4.0 4.5 5.0 5.5 6.0 6.5 7.0

Time [min]

Inte

nsity

1.0

0.8

0.6

0.4

0.3

0.0

0.00 0.25 0.50 0.75 1.00 1.25

Relative concentratiion

Rel

ativ

e re

spon

se

1.0

0.8

0.6

0.4

0.3

0.0

ISTD R

esponse x10 6

1.50 1.75 2.00 2.25 2.50

�� !"#$%&'()*+),-./.012

�� !�� !"#$%&'(�� 3 � 4��

� 3. �� !"#$NBA-AMOZ�� !"# 0.125 g/kg

� 4. �� NBA-AMOZ, �� !"# , 0.125 g/kg-2 g/kg�� !"#$%�� !

5

NBA-d4 AOZ

x104

0.0

0.4

0.8

1.2

NBA-AOZ

x104

0.00

0.50

1.00

1.50

NBA-AHD

10000

20003000400050006000

NBA-d5 AMOZ

x104

0

2

4

6

NBA-AMOZ

0

1

2

3

4

x104Intensity

NBA-SEMx104

01234

2 4 6 8 10 12 Time [min]

�� NBA-AMOZ� NBA-SEM��� !"!#$%

�� !"#$%�NBA-AHD�NBA-AOZ�� !

��� !"#$%&

�� !"#$%&'()*+,-./0 1 µg/kg�

�� !"#$%&'()*+,-./01234

��NBA-AMOZ: 0.125 µg/kg�NBA-AOZ�NBA-SEM:

0.25 µg/kg� NBA-ADH: 0.5 µg/kg��� !"#$

�� !"#$

� 5. �� !"# $%&'( 0.25 g/kg�� !"#$%&'()

6

� 0.125 µg/kg-2 µg/kg�� !�� !"#$%�

�� !"#0.99��� !"#$%&'(�� !

��(RSD)�� 10%��� !"#$ 1��

��

�� XCT�� !"#�� !"#$%&'()*

�� !"#$%&'()*+�� !"#$0.125

µg/kg�0.25 µg/kg�� !"�� !"#$%&'

�� (MRPL)�

�� !

�� !"#$%&'()*+,-.��

www.agilent.com/chem

� 1. �� !"#$%&'()*+,-./()012

NBA-SEM NBA-AOZ NBA-AMOZ NBA-AHDStandard Accuracy % Accuracy % Accuracy % Accuracy %( g/kg) SD % (average) SD % (average) SD % (average) SD % (average)0.125 3.77 98.81 2.29 98.94 4.34 101.10

0.25 2.26 102.72 2.52 101.76 5.87 95.55 6.05 98.31

0.5 3.40 100.72 3.35 100.58 6.21 105.19 4.59 103.87

1 3.56 96.62 3.01 101.90 5.46 99.11 6.53 100.22

2 2.94 101.12 2.13 96.82 5.77 99.05 6.97 97.60

All calibration curves linear weighted 1/x n = 6

7

�� !"#$%&'()*+,-./�� !"�� !"#$%&'()*+,-.

�� !"#$%&'%(

�� !"#$%&�� !"#$

©�� !"#$%&'2004

�� !

2004 � 5 � 11�

5989-0738CHCN

www.agilent.com/chem