Page 66 - Application Notebook - Solution for Food Safety

LAAN-A-LC-E286

Application High Performance Liquid Chromatography

News Analysis of Residual Antimicrobials in Meat with

Antimicrobial Screening System (Part 2)
No.L510

Antimicrobials are a type of veterinary drug and animal Sample (homogenized) 10 g
feed additive, and are used for the treatment and
prevention of disease in livestock and marine products. Extraction
Residual antimicrobials are often found in livestock and
marine products, so threshold levels for antimicrobials Supernatant
are set by regulation to ensure the safety of the Fat Removal
consumer based on amounts that do not harm human
health. Acetonitrile Layer
Due to ongoing reports of recent cases of regulatory Evaporation
violations in various countries and the large number of
compounds targeted for testing, there is a demand for Residue
quick and simple antimicrobial screening.
While Application News No.L509 described an example Redissolution
of using the antimicrobial screening system for Acetonitrile Layer - Aqueous Layer
screening 12 quinolone compounds, this Application
News describes an example screening analysis of 12
antimicrobial target compounds including sulfanomides. HPLC
Fig. 1 Sample Pretreatment Protocol
Q Sample Pretreatment
Sample pretreatment for analysis of residual antimicrobials
in meat usually employs liquid-liquid extraction (and Q Analysis of Antimicrobials Including Sulfonamides
sometimes solid phase extraction), but this process takes in Meat
time and effort. In this article, we employed a QuEChERS Chicken and beef were used as samples. The analytical
method designed to be more efficient and reduce conditions are shown in Table 2. Chromatograms of the
pretreatment times. The QuEChERS method is used to pretreated matrix solutions (blue line), matrix solutions
pretreat vegetables and fruits for residual pesticide spiked with standard solution to create matrix standard
analysis. solutions (red line), and neat standard solution (black
After using the QuEChERS method to perform extraction line) are shown in Fig. 2.
and fat removal, sample solutions were prepared by Standard solution was added to matrix solutions to
evaporation and redissolution steps. Table 1 shows the make up antimicrobial concentrations, including
maximum residue limits (MRLs) of target compounds and sulfonamide concentrations, of 0.01 mg/kg in matrix
sample solution concentrations after sample pretreatment, standard solutions. Standard solutions were prepared to
and Fig. 1 shows the sample pretreatment protocol. Refer the sample solution concentrations listed in Table 1.
to the instruction manual of the system for the details of The photodiode array (PDA) detector (six-wavelength)
the sample pretreatment procedure. built in the i-Series instrument was used for detecting
all target compounds. Employing the analytical
conditions shown, all 12 compounds were separated
Table 1 Maximum Residue Limits and Sample Solution
Concentration of Screening Target Compounds and eluted in approximately 25 minutes.
MRL Sample Solution
Compound Table 2 Analytical Conditions
(mg/kg) Concentration (mg/L)
1 Sulfadiazine 0.01 0.025
System : LC-2040C 3D
2 Sulfamerazine 0.01 0.025 Column : Shim-pack FC-ODS (150 mm L. × 4.6 mm I.D., 3 μm)
3 Sulfadimidine 0.01 0.025 Mobile Phase : A) 20 mM (Sodium) Phosphate Buffer Containing
4 Sulfamonomethoxine 0.01 0.025 0.1 M Sodium Perchlorate
5 Trimethoprim 0.01 0.025 B) Acetonitrile/Methanol=80/20
Time Program : Gradient Elution
6 Sulfamethoxazole 0.01 0.025
Flowrate : 1.0 mL/min
7 Ormetoprim 0.01 0.025 Column Temp. : 50 ˚C
8 Sulfadimethoxine 0.01 0.025 Injection Volume : 20 μL
9 Sulfaquinoxaline 0.01 0.025 Detection : 240 nm
10 Pyrimethamine 0.01 0.025 270 nm
280 nm
11 Difurazon 0.01 0.025
285 nm
12 Nicarbazin *1 0.01 0.025
350 nm
*1: Concentration of N, N'-Bis(4-nitrophenyl)urea, the main 380 nm
Cell Temp. : 40 ˚C
constituent of nicarbazin.