Psilocin and Psilocybin in Urine on Raptor Biphenyl by LC-MS/MS

	Peaks	t_R (min)	Conc. (ng/mL)	Precursor Ion	Product Ion 1	Product Ion 2
1.	Psilocybin	1.36	500	285.1	205.1	240.0
2.	Psilocin-D10	1.58	200	215.1	66.1	-
3.	Psilocin	1.60	500	205.1	160.1	115.0

Column

Raptor Biphenyl (cat.# 9309A52)

Dimensions:

50 mm x 2.1 mm ID

Particle Size:

2.7 µm

Pore Size:

90 Å

Guard Column:

Raptor Biphenyl EXP guard column cartridge 5.0 mm, 2.1 mm ID, 2.7 µm (cat.# 9309A0252)

Temp.:

35 °C

Standard/Sample

Diluent:

Water, 0.1% formic acid + 2 mM ammonium formate

Conc.:

500 ng/mL

Inj. Vol.:

5 µL

Mobile Phase

Water, 0.1% formic acid + 2 mM ammonium formate

Methanol, 0.1% formic acid + 2 mM ammonium formate

Time (min)	Flow (mL/min)	%A	%B
0.00	0.5	95	5
0.20	0.5	95	5
2.50	0.5	5	95
2.51	0.5	95	5
3.50	0.5	95	5

Detector	MS/MS
Ion Source:	Electrospray
Ion Mode:	ESI+
Mode:	MRM
Instrument	HPLC
Sample Preparation	A 500 ng/mL standard mix of psilocin and psilocybin was prepared in pooled urine. A 50 µL aliquot was taken from the standard and mixed with 10 µL of internal standard (psilocin-D10, 20 µg/mL) and 100 µL of methanol. The mixture was vortexed at 3000 rpm for 10 seconds and then centrifuged at 4300 rpm for 10 minutes at 10 °C. After centrifugation, 100 µL of the supernatant was diluted with 900 µL (20-fold dilution) of water containing 0.1% formic acid and 2 mM ammonium formate (mobile phase A) and injected for LC-MS/MS analysis.

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