D e t e r m i n at i o n o f M e l a m i n e R e s i d u e i n Wat e r S a m p l e s by UPL C / MS / MS
Wang Jing, Liu Zhengzheng (Environmental Monitoring Center of Zhejiang Province)
The melamine contamination of milk that occurred in 2008 was
undoubtedly a serious food safety situation in China as safe drinking
water is an important part of food safety. Since both the drinking
water and the food chain may cause a direct or indirect impact on
human health, it is urgent to carry out the study on the analysis
method of melamine residue in water samples.
C h romatog ra p h y Condit ions
UPLC®/MS method: column: UPLC column (ACQUITY UPLC® BEH
HILIC, 1.7 μm, 2.1 50 mm); column temperature: 40 °C; mobile
phase: a mixture of acetonitrile and water (containing 10 mM of
ammonium acetate) (90:10); flow speed: 0.4 ml/min.
The target contaminator is quantitatively analyzed with the MRM
E x p e rim enta l
Equipment and Reagents
An Ultra Performance Liquid Chromatography/tandem mass
spectrometry (ACQUITY®/Quattro Premier™ ); MassLynx™ 4.1
workstation; solid-phase extraction (Zymark); Waters Oasis ®
mode. Detect the target with ESI+. Capillary voltage: 3.0 KV, source
temperature: 120 °C, the temperature of desolvent gas: 400 °C,
flow speed: 800 L/h, cone gas flow: 50 L/h; argon flow speed: 0.38
mL/min at the MRM mode; cone current: 40 V, CID: 17; characteristic
ions pair: 127>85, as in the following Figure 1.
126.8
100
MCX SPE column (200 mg, 6 mL); pressure blowing concentrator
(Organomation).
84.8
Methanol and acetonitrile are both in liquid chromatography pure
(Fisher); water is obtained from a Millipore water purification system;
%
ammonium acetate is in guaranteed grade (Merck), hydrochloric
acid and ammonia are in analytical pure (purchased from East China
Pharmaceutical Company); and the standard reference is purchased
59.8
59.1
66.7
0
55
Sample of the Pretreatment Methods
84.2
67.8
from J&K.
60
65
75.9
83.3
69.2 77.5
70
75
80
86.2
85
94.4 99.0
90
95
100
109.6
109.0 110.2
105
110
115
125.9 128.1
123.0
120
125
130
167.0
176.9 178.6
131.5 140.8 147.2 153.2 158.3 163.8
135
140
145
150
155
160
165
170
175
180
186.6 191.7
185
190
195
200.4
200
202.9
205 m/z
Figure 1. Mass spectra of melamine.
Solid-phase extraction: Add 5 mL of methanol and 5 mL water,
at the speed of 5 mL/min to the column to make it active.
Acidification of water sample: Add 0.25 ml of concentrated
Pe rfo rmance
hydrochloric acid to 500 ml of water. Add the water sample
Accuracy
with the speed of 4 mL/min and the volume of 500 ml. Rinse
Add 6.0 µg of target substance to 500 ml of testing water, and
the sample with 5 mL of water and methanol respectively. Elute
the recovery of a series of pre-sampling is 101.8±10.7% (n=6), as
with 5 mL of methanol (containing 5% of ammonia) twice and
showed in Table 1.
concentrate the eluent with a pressure blowing concentrator
until it is almost dried up. Dilute to 1 mL with mobile phase and
Precision
filter it. Inject.
Inject 1.0 µg/ml of standard solution for 6 times repeatedly and
the RSD of peak area is 3.2%.
Detective limit
0.95
The signal/noise ratio is 15 for 1.0 ng/mL of standard solution
and 3 for 0.2 ng/mL of standard solution, as shown in Figure 2.
Taking the pre-sampling procedure into consideration, the quanti-
%
0.40
0.07
tative detective limit of this method is 0.4 ng/L.
Linearity
6
0.10
Prepare 4 standard solutions with different concentrations ranging
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
1.10
1.20
1.30
1.40
1.50
1.60
1.70
1.80
1.90
2.00 min
Figure 3. TIC of testing sample.
from 1.2 ng/mL to 2400 ng/mL, the equation for the standard
curve is y=1350.3x+161822,R=0.9925.
Conc lusions
Blank
The UPLC/MS/MS method used for the determination of melamine
residue in water sample has been proved to be practicable.
The blank is lower than the detective limit.
The analyses can be fulfilled in 1.0 min with a recovery of
Determination of sample
101.8±10.7% and a precision of 3.2%. With the range covering
The water sample is obtained from a reservoir in Zhejiang province
three magnitudes, the linear coefficient is greater than 0.99 and
with a concentration of melamine of <0.4 ng/L. Add some standard
the detective limit is up to 0.4 ng/L. All of these indicated that
solution to this water sample and analyze the content of melamine.
this method meets the requirement of practical analyses.
The concentration of melamine is 0.03 µg/L and the total ion flow
is showed in Figure 3.
Analytes
Precision
%(n=6)
Recover%
(n=6)
Detective
limit (ng/L)
Linearity
Melamine
3.2
101.8±10.7
0.4
y = 1350.3x
+ 161822,
r=0.9925
Figure 1. Performance of this method.
S/N:PtP=15.12
0.93
0.98
%
0.01
0.130.18
0.40
0.25
0.54
1.05
1.071.13
0.84
0.62 0.690.70
1.24 1.27
1.34 1.41
1.42
1.54
0.32
1.65
1.69 1.77 1.77 1.83 1.90 1.93
3
S/N:PtP=3.48
0.90
0.99
0.00
0.02
0.03
0.07 0.19
%
0.39
0.41
1.01
0.43
0.57 0.67
0.270.29
0.82
0.72 0.79
0.84
1.07
1.17
1.59
1.43 1.46 1.48
1.251.26
1.36
1.63
1.80 1.85
1.71 1.79
1.95 1.98
4
0.10
0.20
0.30
0.40
0.50
0.60
0.70
0.80
0.90
1.00
1.10
1.20
1.30
1.40
Figure 2. The S/N of melamine in different concentration.
1.50
1.60
1.70
1.80
1.90
2.00 min
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