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Silliker

Chem, Res. Center Crete, IL – Report of A Validation of LC-MS/MS Method for Folate Analys

11

Calibration Curve using Dionex 3000 ultimate HPLC and API 5500 MS (Qtrap) detector:

The linearity evaluation of calibration were performed with standards prepared in the elution solvent,

i.e., 10:90:1(v,v,w) :: formic acid : methanol : ascorbic acid and neutralized with ammonium hydroxide.

The calibration curves were extended to 7pts. Concentrations and consisted of: 1, 2, 5, 20, 50, 200 and

500 ng/mL.

Figure 2

:

Calibration Curve of different folate compounds (Dionex 3000 Ultmate HPLC and

5500 API MS detector). The color scheme for the calibration curve of different folate

compounds is as the following

.

Folate (295) ______,

5-CH3-THF (313)_______,

5-CHO-THF

(327)_______,

THF (299) _____,

10-CHO-FA (295)_________,

10-CH3-FA (309) ______.

CH=THF (282) _______. Conc

Calibration Values:

The calibration values based on the above calibration curve for different folate compounds are listed in

the following Table 5.

Table 5: Evaluation of Response Linearity against concentration of six target folate compounds

the values of linear regression

Folate

Compound

Transition

Slope

Intercept

r2

weighing

Calibration

Line Color

Folic Acid

295

0.02807 +0.00269 0.9996

1/x

Navy Blue

5-CH3-THF

313

0.02453 +0.01463 0.9931

1/x

Purple

5-CHO-THF

327

0.02746 +0.01105 0.9922

1/x

Red

THF

299

0.01553 -0.01530

0.9992

1/x

Orange

CH=CH-THF

282

0.1050 +0.00211 0.9965

1/x

Sky Blue

10-CH3-FA

309

0.03783 -0.01387

0.9979

1/x

Light Green

10-CHO-FA

295

0.04435 +0.00650 0.9996

1/x

Dark Green

Folate Compound

Concentration (ppb)

2011.06/Fol-22 (February 2016)

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