Trace Analysis of Salbutamol in Chicken Feed using the

Varian 500 Ion Trap Mass Spectrometer

By Crystal Holt, LC/MS Product Specialist,

Western RegionVarian Customer Service

2

Outline

•Salbutamol Background•Why Salbutamol in chicken feed?•Instrumentation•Method Parameters•Data•Summary•Acknowledgments

3

Salbutamol Background

• Bronchial smooth muscle relaxer

• Asthma medication active ingredient

• Oral or intravenous delivery

• Common side effects include palpitations, muscle cramps and headaches

MW = 239.3

4

Why Salbutamol in Chicken Feed?

• Antibiotics and steroids in feed are being banned in many countries because of disease resistance and human health concerns•Salbutamol is shown to be a viable alternative to antibiotics and steroids • Studies show that salbutamol in chicken feed increases the muscle or lean meat in chickens by over 10%• Studies show that salbutamol in chicken feed increases feed efficiency by over 5%

Note: Studies performed by Stirling Products Limited and Massey University in New Zealand

5

Why Salbutamol in Chicken Feed?

• The world consumes over 46 billion chickens every year• There are 27 chemicals licensed for use in chicken feed which include hormones and antibiotics. The chicken you eat can contain the residue of many toxic chemicals. • Chloramphenicol, an antibiotic was banned for use in Canadian food animals in 1985.

• A suspected human carcinogen it produces leukemia, aplastic anemia and other bone marrow changes.

• When heated to decomposition it emits toxic fumes. • You'd hardly want to enjoy meat containing this

chemical cooking on your barbecue! • In BC alone there are over 40 privately owned and uninspected poultry processing plants.

6

Chicken Feed?

•In Canada in the last decade: • Beef consumption down 16% • Pork consumption down 14% • Poultry consumption up 31%

• In 1994 500 million poultry were slaughtered at federally registered Canadian slaughterhouses. Of these 72.6 million were slaughtered in BC.•The average Canadian eats about 20 birds per year.

7

Performance and productivity through innovation

Enhanced Charge Capacity

Triple Resonance Mass Scanning

Superior Sensitivity

MS10

SelecTemp

Varian 500 Ion Trap MS

8

Enhanced Charge Capacity-ECCTM

Mechanical & electrical modification of multipole content of trapping field guide

• Stores more ions before affects on mass resolution and mass position

• X10 increase in charge capacity compared to common 3D traps

• Storing the same number of analyte ions and X10 more matrix ions without space charging effects

Features & Benefits

9

Charge capacity effects

X10

Peak width vs. ion time Varian 500-MS and Competitors 3D Trap

Varian 500-MSCompetitor

Equals 10X charge capacity

Proof – 10X ion time

10

Charge capacity effects

X10

Varian 500-MSCompetitors

Mass shift vs. ion time Varian 500-MS and Competitors 3D Trap

Equals 10X charge capacity

Proof – 10X ion time

11

Method Parameters

500 Ion Trap Parameters

Mode: ESI (+)Source Temp: 65 degreesDry Gas Temp: 350 degreesCapillary Voltage: 66RF %: 72%Needle Voltage:4500CID: 0.61 (240-222), 2.80 (222-148), 1.80 (240-222-148)

HPLC conditions

Column: Pursuit C18, 3 micron, 150X3mmFlow Rate: 0.4 ml/min, isocraticMobile Phase: 75% water, 24% acetonitrile, 1% formic acidInjection Volume: 20 ul

12

Signal/Noise Ratio Comparison of MS/MS Scan vs MS/MS/MS Scan

S/N: 4099

S/N: 4674

S/N: 884

13

MS/MS/MS Chromatogram (240-222-148)

14

Salbutamol Calibration Curve

Calibration Range: 0.5ppb to 100ppb; r2 = 0.9998

15

Salbutamol Spikes Sample (1.3ppb)

16

Limit of Detection of Salbutamol (0.01ppb)

17

Summary

• The comparison chromatography test shows that MSn (MS3 for this study) has higher signal/noise ratio than MS/MS scans • Salbutamol had linear response with an r2 of 0.9998 with the Varian 500 Ion Trap Mass Spectrometer• The limit of detection is 0.01ppb having a signal-to-noise ratio of 10:1• Varian 500 Ion Trap Mass Spectrometer is well suited for trace analysis and quantification of salbutamol in chicken feed

18

Acknowledgments

• Eugene Chang, Varian Field Chemist • Thailand Department of Livestock Development• WCAT

Thank you for your attention

Questions?

19

20

Varian 1200L LC/MS

ProStar Binary 210Solvent Delivery

System

ProStar 430AutoSampler

Module

1200L MS with ESI