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LC-MS ANALYSIS OF STEVIA EXTRACT ON HALO-5 PENTA-HILIC

PostPosted: Fri 23 Jun 2017 14:00
by Interchim
LC-MS ANALYSIS OF STEVIA EXTRACT ON HALO-5 PENTA-HILIC
HALO by AMT
Application n°124-F

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TEST CONDITIONS:
Column: 3.0 x 250 mm, HALO-5 Penta-HILIC
Part Number: 95813-905
Mobile Phase:
A= 50/50 Water/acetonitrile with 5 mM
Ammonium formate, pH 3
B= 5/95 Water/acetonitrile with 5 mM
Ammonium formate, pH 3
Gradient : 90% B to 67% B over 30 min
Flow Rate: 0.5 mL/min.
Pressure: 60 bar
Temperature: Ambient
Injection Volume: 5 μL
Sample Solvent: 80/20 Acetonitrile/water
LC System: Shimadzu Nexera
MS: Shimadzu LCMS 2020(single quadrupole)
ESI: +4.5 kV,
Scan range: 200-1200 m/z
Scan rate: 2 pps
Capillary: 250 ˚C
Heat block: 350 ˚C
Nebulizing gas flow: 1.5 L/min
Drying gas flow: 15 L/min

Extraction Procedure
1. Weigh 400 mg of Stevia rebaudiana leaves (Sigma S5381)
2. Crush leaves with mortar and pestle and transfer to vial
3. Add 8 mL of 50/50 (v/v) acetonitrile/water
4. Sonicate vial contents for 15 minutes
5. Filter sample using 25 mm syringe filter having 0.2 μm PTFE membrane (VWR 28145-495)
6. Centrifuge @ 10K rpm (5 min.) and collect supernate
7. Dilute 400 μL of extract in 600 μL of acetonitrile for overall concentration of 80/20 acetonitrile/water
8. Centrifuge diluted sample @ 10K (5 min.) rpm and inject the supernate

Stevia is a natural sweetener and is used as a substitute for sugar. LC/MS analysis of stevia glycosides from a Stevia extract is easily accomplished using HALO-5 Penta-HILIC column due to its unique bonded phase containing five OH groups and the high efficiency of the 5-micron Fused-Core particles.

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