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Chiral Separation of Dextromethorphan/Levomethorphan and Dextrorphan/Levorphanol on Lux 3u AMP, 150x4.6mm

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Sample Preparation Details

Part Number :8E-S029-TGB

Description :Strata™ X-C 33 µm Polymeric Strong Cation

Condition 1 :1 mL methanol

Sample Pretreatment :Urine sample pretreatment: Combine 200µL urine sample, 200µL 100mM ammonium acetate buffer (pH 4.0), 20µL internal standard solution (250ng/mL), and 20µL of beta-glucuronidase (>100,000 units/mL) and incubate for 1 hour at 55°C. Centrifuge for 10 minutes at 13,000rcf. Dilute 400µL supernatant 1:1 with 0.1% aqueous formic acid. Serum sample pretreatment: Combine 250µL serum, 250µL 0.1% formic acid, and 20µL internal standard solution (250 ng/mL).

Load :Full pretreated sample (from above)

Wash 1 :1.0% formic acid in water

Wash 2 :1.0% formic acid in methanol

Dry :3-4 minutes at high vacuum

Elution 1 :2 x 0.5 mL 5% NH4OH in 50:50 methanol/acetonitrile

Evaporate: :Evaporate to dryness

Reconstitute :100 µL 40:60 acetonitrile/water

Condition	1 mL methanol
Condition	1 mL methanol
Wash	1.0% formic acid in methanol
Wash	1.0% formic acid in water
Wash	1.0% formic acid in methanol
Dry	3-4 minutes at high vacuum
Elute	2 x 0.5 mL 5% NH4OH in 50:50 methanol/acetonitrile
Dry Down	at 40°C under a gentle stream of nitrogen
Reconstitute	100 µL 40:60 acetonitrile/water

Application Detail (App ID: 25782)

Column: Lux® 3 µm AMP, LC Column 150 x 4.6 mm, Ea

Part No:00F-4751-E0

Dimensions:150 x 4.6 mm ID

Elution Type:Gradient

Elution A:5mM ammonium bicarbonate, adjusted to pH 11 with ammonium hydroxide

Elution B:Acetonitrile

Gradient Profile:

Step No.	Time (min)	Pct A	Pct B
1	0	55	45
2	4.5	55	45
3	4.6	20	80
4	8	20	80
5	8.1	55	45
6	10	55	45

Flow Rate:1 mL/min

Col. Temp: 40 °C

Detection: LC/MS/MS @ (0°C)

Note:Urine hydrolyzed with beta-glucuronidase and extracted with Strata-X-C

Analyte Details

Analyte Detail not available

Dextrorphan-D3

Dextrorphan 1

Dextrorphan 2

Levorphanol 1

Levorphanol 2

Dextromethorphan-D3

Dextromethorphan 1

Dextromethorphan 2

Levomethorphan 1

Levomethorphan 2