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Simultaneous Determination of Choline/Carnitine in Infant Formulas and Adult Nutritional
Products by HILIC LC-MS/MS
Wei Jing, Lou Salvati, and Joseph J. Thompson, Abbott Nutrition, Columbus, Ohio USA
Principle
The reconstituted test sample is weighed into a microwave reaction vessel. Microwave heating
accelerates an acidic hydrolysis process to release bound choline. A subsequent alkaline degradation is
performed to release L-carnitine from inherent acylcarnitines. Choline and L-carnitine can be determined
quantitatively in nutritional products and raw materials by hydrophilic interaction liquid chromatography
with tandem mass spectrometry (HILIC LC-MS/MS).
A.
Apparatus
a.
Column
- Acquity UPLC BEH HILIC 1.7
m, 2.1×100 mm (Waters Corp., Milford, MA)
or equivalent
b.
Liquid chromatograph
—Waters Acquity UPLC Binary
c.
Solvent Manager
- capable of 15 000 psi or equivalent
d.
Detector
—Waters Quattro Premier XE, or Xevo TQ-S triple quadrupole mass
spectrometer, or equivalent
e.
Injector
—Waters Acquity sample manager with integrated column oven or equivalent
f.
Nitrogen generator
.—Peak Scientific (Billerica, MA) Model NM30LA or equivalent
g.
Data system
—Waters MassLynx, latest revision or equivalent
h.
Microwave
– MarsXpress (CEM, Matthews, NC) or equivalent
i.
Reaction Vessel
- 50 mL TEFL reaction vessels (CEM, Matthews, NC)
j.
Centrifuge tubes
— PP (Polypropylene), 50 mL capacity disposable or equivalent
k.
Vortex mixer with flat and cone top
l.
Balance1 —
Readable to at least 0.0001 g. ( Mettler-Toledo AT200 or equivalent)
m.
Balance2 —
Readable to at least 0.001mg. (Mettler-Toledo XP 6 or equivalent)
n.
Syringe –
1 mL Luer tip
o.
Filters
– 0.45 µm Nylon membrane with syringe tip or equivalent
p.
Pipets
- 5~25, 50~250, and 100~1000 μL adjustable
q.
Pipet tips
B.
Reagents
a.
Solvents and Chemicals
i.
Acetonitrile
—Fisher Optimal LCMS grade, or equivalent
ii.
Nitric Acid
— 69-70% Reagent grade.
J.T. Baker or equivalent
iii.
Ammonium Acetate
—Fluka LCMS additive grade or equivalent
iv.
Ammonium Hydroxide
—28 – 30% Reagent grade (for adjusting pH in mobile
phase also, check it before use). MACRON or equivalent.
v.
Distilled or deionized laboratory water
—Purified water, USP; ≥18Mohm/cm.
Candidates for 2016 Method of the Year
325