Theanine by HPLC-ELSD
Determination of Theanine by HPLC-ELSD
1) Reagents
a. Nanopure water b. Acetonitrile (HPLC) c.TFA(C.R)
2) Apparatus
HPLC (Shimadzu) Analytical balance (1/10000) Ultrasonic cleaner
0.45μm PTFE filter 50ml volumetric flask Screw cap vials
ELSD detector(Alltech)
3) HPLC condition
Column: Polaris C18-A 5 m 4.6250mm
Mobile Phase: A:0.1% TFA of Water B: 0.1% TFA of Acetonitrile
Flow Rate: 0.5ml/min
Injection: 10μl
Drift tube temperature :95℃
Gas flow:2.5L/min
Impactor: Off
Gain:2
Gradient:
Time %A %B
0.01 100 0
5 100 0
20 60 40
25 60 40
28 100 0
30 100 0
4) Procedure
Samples Preparation
Accurately weigh 45mg extraction into 50ml volumetric flask. Add Water to volume and sonicate to mix, filter through a 0.45μm PTFE filter into a screw cap vial and proceed with the analysis.
Standard preparation /calibration
Prepare a standard solution (approximately 0.2mg/ml in Water using a standard of Theanine，Sonicate to mix, filter through a 0.45μm PTFE filter into a screw cap vial and proceed with the analysis.
5) Calculation
Determine the peak area’s logarithm of Theanine the on the sample chromatogram and compare to the corresponding peaks in the chromatogram for the standard solution.
1) Reagents
a. Nanopure water b. Acetonitrile (HPLC) c.TFA(C.R)
2) Apparatus
HPLC (Shimadzu) Analytical balance (1/10000) Ultrasonic cleaner
0.45μm PTFE filter 50ml volumetric flask Screw cap vials
ELSD detector(Alltech)
3) HPLC condition
Column: Polaris C18-A 5 m 4.6250mm
Mobile Phase: A:0.1% TFA of Water B: 0.1% TFA of Acetonitrile
Flow Rate: 0.5ml/min
Injection: 10μl
Drift tube temperature :95℃
Gas flow:2.5L/min
Impactor: Off
Gain:2
Gradient:
Time %A %B
0.01 100 0
5 100 0
20 60 40
25 60 40
28 100 0
30 100 0
4) Procedure
Samples Preparation
Accurately weigh 45mg extraction into 50ml volumetric flask. Add Water to volume and sonicate to mix, filter through a 0.45μm PTFE filter into a screw cap vial and proceed with the analysis.
Standard preparation /calibration
Prepare a standard solution (approximately 0.2mg/ml in Water using a standard of Theanine，Sonicate to mix, filter through a 0.45μm PTFE filter into a screw cap vial and proceed with the analysis.
5) Calculation
Determine the peak area’s logarithm of Theanine the on the sample chromatogram and compare to the corresponding peaks in the chromatogram for the standard solution.