Applications

TMP and Thiamine Standard by Impact PPT on Kinetex 5u C18 100x4.6mm

Analytes

1Thiamine monophosphate
2Thiamine

Compound Name Monofosfotiamina
CID 10761
Molecular Formula C12H18ClN4O4PS
Molecular Weight 380
No. Hydrogen Bond Acceptors 8
No. Hydrogen Bond Donors 3

Details

LC Conditions (App ID: 20875)

Column:

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Brand Name:

Kinetex

Part No:

00D-4601-E0

Phase Name:

C18

Elution Type:

Gradient

Mobile Phase:

A: 25mM Na2HPO4, 10% Methanol, pH 7.0

B: 25mM Na2HPO4, 70% Methanol, pH 7.0

Gradient Profile:

Step No.	Time(min)	%A	%B
1	0	97	3
2	0.25	75	25
3	0.75	75	25
4	3	65	35
5	4	40	60
6	4.5	97	3
7	5	97	3

Flow Rate:

1 mL/min

Temperature:

25°C

Detection:

Fluorescence (FLUOR) @ Ex. 375 nm (22°C)

Sample Note:

Using Impact Protein Precipitation Plate (Cat Log #CE0-7565) for sample preparation

Sample Preparation Details

Description: Impact Protein Precipitation, 2mL Square Well Filter Plate, 2/Pk
Part Number: CE0-7565
Pre Treatment Note: Add 400µL of methanol to a well of a Protein Precipitation Plate; Add 100µL plasma to the well. Mix 3 times with pipette tip (or vortex the whole plate briefly); Wait for 5 minutes; Filter through the Protein Precipitation Plate under vacuum (5 Hg) for 5 minutes. Transfer 50 µL to an autosampler vial (or a well of 96-well plate). Add 100µL water and 25 µL of derivatizing reagent. Cover the vial with a lid. Vortex for 15 seconds. Put the vial into an autosampler. All conditions are at room temperature (RT). It is ready to inject. Derivatizing reagent: Potassium ferricyanide [K3Fe(CN)6] was prepared as the following: dissolve 6mg of K3Fe(CN)6 in 15mL 15% sodium hydroxide solution. Prepare fresh daily.
Sample PreTreatment: Add 400µL of methanol to a well of a Protein Precipitation Plate; Add 100µL plasma to the well. Mix 3 times with pipette tip (or vortex the whole plate briefly); Wait for 5 minutes; Filter through the Protein Precipitation Plate under vacuum (5 Hg) for 5 minutes. Transfer 50 µL to an autosampler vial (or a well of 96-well plate). Add 100µL water and 25 µL of derivatizing reagent. Cover the vial with a lid. Vortex for 15 seconds. Put the vial into an autosampler. All conditions are at room temperature (RT). It is ready to inject. Derivatizing reagent: Potassium ferricyanide [K3Fe(CN)6] was prepared as the following: dissolve 6mg of K3Fe(CN)6 in 15mL 15% sodium hydroxide solution. Prepare fresh daily.
Evaporation: Evaporate to dryness

Order Items Used in This Application

00D-4601-E0

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Qty:

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TMP and Thiamine Multi-Standard by Impact PPT on Kinetex 5u C18 100x4.6mm

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Reversed Phase

TMP and Thiamine Standard 5 nmol/L by Impact PPT on Kinetex 5u C18 100x4.6mm

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Reversed Phase

TMP and Thiamine Standard by Impact PPT on Kinetex 5u C18 100x4.6mm

Order Items Used in This Application

00D-4601-E0

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Details

LC Conditions

Column:

Kinetex 5 µm C18 100 Å, LC Column 100 x 4.6 mm, Ea

Brand Name:

Kinetex

Part No:

00D-4601-E0

Phase Name:

C18

Elution Type:

Gradient

Mobile Phase:

A: 25mM Na2HPO4, 10% Methanol, pH 7.0
B: 25mM Na2HPO4, 70% Methanol, pH 7.0

Gradient Profile:

Step No.	Time(min)	%A	%B
1	0	97	3
2	0.25	75	25
3	0.75	75	25
4	3	65	35
5	4	40	60
6	4.5	97	3
7	5	97	3

Flow Rate:

1 mL/min

Temperature:

25°C

Detection:

Fluorescence (FLUOR) @ Ex. 375 nm (22°C)

Sample Note:

Using Impact Protein Precipitation Plate (Cat Log #CE0-7565) for sample preparation

LC Sample Preparation Details

Description:	Impact Protein Precipitation, 2mL Square Well Filter Plate, 2/Pk
Part Number:	CE0-7565
Phase Name:	Protein Precipitation (PPT)
Pre Treatment Note:	Add 400µL of methanol to a well of a Protein Precipitation Plate; Add 100µL plasma to the well. Mix 3 times with pipette tip (or vortex the whole plate briefly); Wait for 5 minutes; Filter through the Protein Precipitation Plate under vacuum (5 Hg) for 5 minutes. Transfer 50 µL to an autosampler vial (or a well of 96-well plate). Add 100µL water and 25 µL of derivatizing reagent. Cover the vial with a lid. Vortex for 15 seconds. Put the vial into an autosampler. All conditions are at room temperature (RT). It is ready to inject. Derivatizing reagent: Potassium ferricyanide [K3Fe(CN)6] was prepared as the following: dissolve 6mg of K3Fe(CN)6 in 15mL 15% sodium hydroxide solution. Prepare fresh daily.
SPE Sample PreTreatment:	Add 400µL of methanol to a well of a Protein Precipitation Plate; Add 100µL plasma to the well. Mix 3 times with pipette tip (or vortex the whole plate briefly); Wait for 5 minutes; Filter through the Protein Precipitation Plate under vacuum (5 Hg) for 5 minutes. Transfer 50 µL to an autosampler vial (or a well of 96-well plate). Add 100µL water and 25 µL of derivatizing reagent. Cover the vial with a lid. Vortex for 15 seconds. Put the vial into an autosampler. All conditions are at room temperature (RT). It is ready to inject. Derivatizing reagent: Potassium ferricyanide [K3Fe(CN)6] was prepared as the following: dissolve 6mg of K3Fe(CN)6 in 15mL 15% sodium hydroxide solution. Prepare fresh daily.
SPE Evaporation:	Evaporate to dryness

Analytes

1. Thiamine monophosphate

2. Thiamine

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