LC Technical Note

Simultaneous Analysis of Cyanoguanidine and Melamine
in Food by LC/MS/MSLT132
GL Sciences Inc.

It became a serious problem in China and Taiwan in2013 that cyanoguanidine , or dicyandiamide, wasdetected in powdered milk made in New Zealand. Since
cyanoguanidine is contained in some chemical fertilizer
as nitrification inhibitor, it is considered as a possible
cause of the problem that such fertilizer was used inranch.
In 2008, melamine was detected in milk and dairy
products, which also became a big issue. HILIC(hydrophilic interaction chromatography) mode was used
for the determination of melamine content
because melamine is highly hydrophilic.
Chemical structure of cyanoguanidine is similar to that ofmelamine. In this note, cyanoguanidine was analyzed withHILIC mode. Cyanoguanidine and melamine could bedetermined simultaneously using Inertsil HILIC as aseparation column.
(K.Kanno)

Chromatogram Obtained from Standard Sample
7.0e4(20 µg/L)(10 µg/L)

1. Cyanoguanidine
2. Melamine

6.0e4
2Intensity [ cps]

5.0e4
4.0e4
3.0e4
1

2.0e4

Q1/Q3 ： 85.1/68.11.0e4

Q1/Q3 ： 126.9/68.1
0.00123456Time [min]78

Flow Rate
Col. Temp.
Detection
Inj. Vol.

Cyanoguanidine: LC800: Inertsil HILIC(5 μm, 150 x 3.0 mm I.D.): A) CH3CN
B) 10 mM Ammonium acetate
A/B = 90/10 － 0.5 min － 90/10 － 5.5 min －50/50 (Eqilibration for 5 min), v/v：0.5 mL/min:40 ℃

Melamine

1600000
Peak area

Eluent10

2000000

HPLC Condition
SystemColumn

9R2=0.997

1200000
800000: MRM (SRM): 5.0 µL

400000R2=0.998

MS/MS（MRM）Condition
SystemDetection: 4000 QTRAP (ABSciex): Compound
Cyanoguanidine
Melamine

Ion Source: ESI （Posi）

CUR
10Q1
85.1
126.9
CAD8Q368.1
68.1
IS5500

0DP4141

CE4141CXP
610EP10 010

TEM700

GS160

GS230

iheon

510152025
Concenteration [µg/L]
Calibration Curve

303540GL Sciences LC Technical Note

Chromatograms Obtained from Powdered Milk Sample

Example ofSample Pretreatment

Q1/Q3 ： 85.1/68.1

1.5e4

Intensity [cps]Sample— 1.0 gExtraction

1.
2.

Cyanoguanidine
Melamine

11.0e45000— 20 mL of water/acetonitrile= 50/50 (v/v)

Standardaddition
Without addition— Sonication extraction for 30min

00 1 2 3 4 5— Filtration (0.45 µmmembrane filter)6 7 8 9Time [min]10Q1/Q3 ： 126.9/68.1

HPLC

2Intensity [cps]1.5e4

1.0e45000
Standardaddition
Without addition

00 1 2 3 4 5

6 7 8 9Time [min]10

Cyanoguanidine and melamine was added to commercially available powdered milk.
Each concentration was adjusted to 10 µg/L ( = 0.2 mg/kg powdered milk）.

< HPLC Column used in This Note>
Inertsil HILIC 5 μm, 150 x 3.0 mm I.D. (Cat.No. 5020-07715)

GL Sciences disclaims any and all responsibility for any injury or damage which may be caused by this data directly or indirectly. We reserve theright to amend this information or data at any time and without any prior announcement.

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