25 ng/mL Benzodiazepines in Whole Blood on Ultra Biphenyl by LC/MS/MS

	Peaks	t_R (min)	MRM1	MRM2	MRM3
1.	7-Aminoclonazepam	0.57	286.0/250.3	286.0/222.2	286.0/121.0
2.	7-Aminoflunitrazepam	0.74	284.2/135.2	284.2/240.2	284.2/226.1
3.	Bromazepam	1.07	315.9/182.0	315.9/209.1	315.9/260.9
4.	N-Desmethylflunitrazepam	1.70	300.0/254.1	300.0/225.2	300.0/198.0
5.	Nitrazepam	1.68	282.1/236.1	282.1/207.2	282.1/180.0
6.	Lorazepam	1.70	321.0/302.9	321.0/275.1	321.0/229.1
7.	2-Hydroxyethylflurazepam	1.78	333.1/246.1	333.1/166.0	333.1/109.1
8.	Nordiazepam	1.76	271.0/140.0	271.0/208.0	271.0/225.9
9.	Estazolam	1.86	294.9/267.1	294.9/241.0	294.9/138.0
10.	Desalkylflurazepam	1.90	289.0/140.0	289.0/165.1	289.0/226.0

	Peaks	t_R (min)	MRM1	MRM2	MRM3
11.	Clonazepam	1.91	315.9/270.2	315.9/214.1	315.9/207.1
12.	Alprazolam	1.98	309.4/281.1	309.4/205.0	309.4/274.1
13.	Midazolam	1.22	326.1/291.2	326.1/244.2	326.1/209.0
14.	Triazolam	2.05	343.1/315.0	343.1/308.2	343.1/239.2
15.	Temazepam	2.12	301.2/283.1	301.2/255.1	301.2/228.1
16.	Flurazepam	1.30	388.1/315.1	388.1/288.2	388.1/317.1
17.	Flunitrazepam	2.23	314.4/211.0	314.4/239.1	314.4/268.1
18.	Clobazam	2.26	301.1/259.1	301.1/224.0	301.1/181.1
19.	Diazepam-D5 (IS)	2.40	290.0/154.1	--	--
20.	Diazepam	2.44	284.8/193.1	284.8/222.2	284.8/257.1

Column

Ultra Biphenyl (cat.# 9109552)

Dimensions:

50 mm x 2.1 mm ID

Particle Size:

5 µm

Pore Size:

100 Å

Temp.:

40 °C

Standard/Sample

Diluent:

Starting mobile phase + 30 ng/mL IS

Conc.:

25 ng/mL sample was diluted 20x prior to injection

Inj. Vol.:

30 µL

Mobile Phase

Water + 2 mM ammonium formate + 0.2% formic acid

Acetonitrile + 10% water + 2 mM ammonium formate + 0.2% formic acid

Time (min)	Flow (mL/min)	%A	%B
0.00	0.6	70	30
3.00	0.6	40	60
3.1	0.6	5	95
3.5	0.6	5	95
3.6	0.6	70	30
4.5	stop

Detector	AB SCIEX API 4000 MS/MS
Model #:	API 4000
Ion Source:	TurboIonSpray^®
Ion Mode:	ESI+
Ion Spray Voltage:	2 kV
Curtain Gas:	40 psi (275.8 kPa)
Gas 1:	40 psi (275.8 kPa)
Gas 2:	40 psi (275.8 kPa)
Source Temp.:	600 °C
Mode:	MRM
Instrument	Applied Biosystems/MDS Sciex LC-MS/MS System
Notes	MS/MS instrument was operated in scheduled MRM mode.

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