25-OH Vitamin D2 and D3 Control from Serum on Kinetex 2.6µm C18 after PPT

LC Conditions

Column

Kinetex 2.6 µm C18 100 Å, LC Column 50 x 4.6 mm, Ea

Brand

Kinetex

Part No

00B-4462-E0

Phase Name

C18

Elution Type

Gradient

Mobile Phase

A: 0.05 % Formic acid

B: 5 mM Ammonium acetate + 0.1 % Formic acid in methanol

Gradient Profile

Step No.

Time(min)

%A

%B

1

0

92

8

2

0.08

92

8

3

3.4

0

100

4

4.8

0

100

5

6

92

8

Flow Rate

1.00 mL/min

Temperature

0°C

Detection

Mass Spectrometer (MS) @ (22°C)

Detection Info

Sample Preparation Details

Description

Strata-X 33 µm Polymeric Reversed Phase, 30 mg / 3 mL, Tubes, 50/Pk

Part No

8B-S100-TBJ

Pretreatment Note

Note: Avoid over drying eluates to avoid loss of analytes and/or internal standard, which will impact both precision and accuracy of data Reconstitute with 100 µL ethyl acetate/acetonitrile (5:95), and vortex mix for 5 seconds Centrifuge for 10 min at 2000g to settle any precipitate Transfer clear liquid to amber-colored autosampler vial and cap

Condition 1

1 mL Acetonitrile

Condition 2

1 mL Acetonitrile/Water (65:35)

Load

Load 1.60 mL Sample diluted in Acetonitrile/Water (50:50)

Wash 1

0.5 mL Acetonitrile/Water (65:35) at 1 mL/min

Wash 2

0.5 mL Acetonitrile/Water (65:35) at 1 mL/min

Elution 1

0.5 mL Acetonitrile at 1 mL/min

Elution 2

0.5 mL Acetonitrile at 1 mL/min

Evaporation

Evaporate to dryness

Reconstitute

100 µL ethyl acetate/acetonitrile (5:95)