Extraction of Mono Hydroxy Vitamin D2 and D3 from Phree on Kinetex C18 2.6u 30x3 mm

LC Conditions

Column

Kinetex 2.6 µm C18 100 Å, LC Column 30 x 3 mm, Ea

Brand

Kinetex

Part No

00A-4462-Y0

Phase Name

C18

Elution Type

Gradient

Mobile Phase

A: 0.1% formic acid/water

B: 0.1% Formic acid/methanol

Gradient Profile

Step No.

Time(min)

%A

%B

1

0

40

60

2

0.5

5

95

3

2

5

95

4

2.01

40

60

5

3.5

40

60

Flow Rate

0.60 mL/min

Temperature

22°C

Detection

Mass Spectrometer (MS) @ (22°C)

Detection Info

Sample Notes

Instrument: API 5000 (AB SCIEX) Ionization Source: APCI, Positive Polarity Ionization Source Parameters Curtain Gas: 25.0 psi Gas 1 (Neb Gas): 40.00 psi ihe: ON CAD (Collision Gas): 10.0 NC: 5.00 µA TEM: 360 °C Note: Gas 2 is not available in APCI source MRM Transitions and Mass Dependent Parameters: Compound Q1, Da Q3, Da Dwell, msec DP, V EP, V CE, V CXP, V Vit D2 (1) 395.3 209.3 45 69 10 25 10 Vit D2 (2) 395.3 269.2 45 69 10 25 10 IS (D3-3H2) 386.2 257.2 45 69 10 25 10 Vit D3 (1) 383.2 257.2 45 69 10 23 10 Vit D3 (2) 383.2 211.1 45 69 10 34 10

Sample Preparation Details

Description

Phree Phospholipid Removal, 30 mg / well, 96-Well Plates, 2/Pk

Part No

8E-S133-TGB

Load

Load Sample

Evaporation

Evaporate to dryness

Reconstitute

0 µL initial mobile phase