Determination of melamine in raw milk and dairy products (1)

The General Administration of Quality Supervision, Inspection and Quarantine of the People’s Republic of China, in the "Detailed Rules for the Examination of Licensing Conditions for the Production of Infant Formula Milk Powder by Enterprises (2010 Edition)", for the first time requires companies to conduct melamine inspections on batches of raw milk and raw milk powder purchased.
It also requires related companies to have at least two inspectors who can independently inspect melamine
.

Regarding the determination of melamine , China issued the "GB/T22388-2008 Method for Testing Melamine in Raw Milk and Dairy Products " and "GB/T22400-2008 Rapid Detection of Raw Milk Melamine" in October and November 2008, respectively
.

This training uses high performance liquid chromatography with a limit of quantification of 2 mg/kg (according to the first method of GB/T22388-2008)
.

1.
Purpose requirements

(1) Understand the principle of high performance liquid chromatography for the determination of melamine in raw milk and dairy products
.

(2) Master the method and operation of the establishment of high performance liquid chromatograph instrument conditions
.

2.
Principles of training

The melamine in the sample was extracted with trichloroacetic acid solution- acetonitrile, purified by a cation exchange solid phase extraction column, and determined by high performance liquid chromatography, and quantified by an external standard method
.

3.
Training supplies

(1) High performance liquid chromatograph: Agilent 1100 is equipped with ultraviolet detector or diode array detector
.

(2) Analytical balance: Sensitivity is 0.
0001g and 0.
01g
.

(3) Centrifuge: the speed is not less than 4000r/min
.

(4) Ultrasonic water bath
.

(5) Solid phase extraction device
.

(6) Water bath nitrogen drying instrument: equipped with nitrogen cylinder or other nitrogen source
.

(7) Vortex mixer
.

(8) Plastic centrifuge tube with stopper: 50mL
.

(9) Research body
.

(10) Mixed-type cation exchange solid phase extraction column: Specification 60mg 3mL, it is recommended to use MCX from WATERS or PCX from Bona-Agel or equivalent
.

(11) Methanol: chromatographically pure
.

(12) Acetonitrile: chromatographically pure
.

(13) Ammonia water: the content is 25%-28%
.

(14) Trichloroacetic acid
.

(15) Citric acid
.

(16) Sodium octane sulfonate : chromatographically pure
.

(17) Aqueous methanol solution: accurately measure 50 mL methanol and 50 mL water, mix well and set aside
.

(18) Trichloroacetic acid solution (1%): accurately weigh 10g of trichloroacetic acid into a 1L volumetric flask, dissolve it with water and dilute to the mark, mix well and set aside
.

(19) Ion-pair reagent buffer: accurately weigh 2.
10g of citric acid and 2.
16g of sodium octanesulfonate , add about 980mL of water to dissolve, adjust the pH to 3.
0, and dilute to 1L for later use
.

(20) Ammoniated methanol solution (5%): Accurately measure 5mL ammonia and 95mL methanol, mix well and set aside
.

(21) Melamine standard product: CAS108-78-01, purity greater than 99.
0%
.

(22) Melamine standard stock solution: accurately weigh 100mg (accurate to 0.
1mg) of melamine standard substance in a 100mL volumetric flask, dissolve it with methanol aqueous solution and dilute to the mark, prepare a standard stock solution with a concentration of 1mg/mL, Store at 4°C in the dark
.

(23) trichloroacetic acid solution saturated with n-hexane (optional): Take 200mL hexane and 100mL of trichloroacetic acid into the solution with 500mL separatory funnel, mixed thoroughly shaken, standing layer, the upper layer of n-hexane standby
.

Unless otherwise specified, all reagents used are of analytical grade
.

4.

(1) Trichloroacetic acid has inhalation toxicity and corrosiveness, so it needs to be used and configured in a fume hood, and rubber gloves should be used for operation

(2) Methanol is highly toxic.

(3) The samples must be placed symmetrically during the operation of the centrifuge.

Related links: Detection of illegal additives and other harmful substances in food