Extraction of 5-Fluorouracil from Human Serum Using Strata-X PRO SPE on a Kinetex 2.6 μm PS C18 (ESI -Ve Mode)

LC Conditions

Column

Kinetex 2.6 µm PS C18 100 Å, LC Column 150 x 3 mm, Ea

Brand

Kinetex

Part No

00F-4780-Y0

Phase Name

PS C18

Elution Type

Gradient

Mobile Phase

A: 0.1% Formic acid in Water

B: Methanol

Gradient Profile

Step No.

Time(min)

%A

%B

1

0

93

7

2

12

93

7

Flow Rate

0.20 mL/min

Temperature

25°C

System

Agilent Technologies 1260 Infinity SCIEX 4500

Detection

LC/MS/MS @ (25°C)

Detection Info

Sample Notes

Add 100 µL Serum (doubly stripped) to tube. Add 900 µL of chilled (~00C) Acetonitrile to the serum while vortexing. Centrifuge samples at 6000 rpm for 5 minutes and transfer supernatant.

Sample Preparation Details

Description

Strata-X PRO, 96-Well Plate, 30mg/well, Ea

Part No

8E-S536-TGA

Condition 1

1 mL of ACN

Sample Pretreatment

Sample Pre-treatment: • Add 100 µL Serum to tube • Add 900 µL of chilled (~0°C) ACN to the serum while vortexing • Centrifuge samples at 6000 rpm for 5 minutes and transfer supernatant

Load

supernatant (collected from pre-treatment)

Evaporation

Evaporate to dryness

Reconstitute

100 µL of initial mobile phase

Steps

0

Condition

1 mL of ACN

1

Load

supernatant (collected from pre-treatment)

2

Vacuum

Apply and collect extract

3

Dry Down

Sample under N2 and heat at 45 °C

4

Reconstitute

100 µL of initial mobile phase