NBD-F (4-Fluoro-7-nitrobenzofurazan) is a pro-fluorescent reagent which is developed for amino acid analysis. NBD-F reacts with primary or secondary amines to produce a fluorescent product and used for analysis of amino acids and low molecular weight amines^[1].

As the percentage of the organic phase changes, the retention time of NBD-F remains relatively stable, while the retention times of the derivatization products changes. The pH of the mobile phase affects the separation of the NBD-F and the derivatization products^[1]. NBD-F is a fluorescent derivatization reagent that is originally developed for amino acid analysis, and recently applied to the analysis of other amino acid derivatives such as N-methyl-D-aspartic acid and glutathione. The use of NBD-F appears to have several advantages in that the derivatization procedure is simple and its derivatives are highly stable^[2].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

183.10

C₆H₂FN₃O₃

29270-56-2

4-氟-7-硝基-2,1,3-苯并氧杂噁二唑

Room temperature in continental US; may vary elsewhere.

4deg;C, protect from light

*In solvent : -80deg;C, 6 months; -20deg;C, 1 month (protect from light)

DMSO : 100 mg/mL (546.15 mM; Need ultrasonic)

请根据产品在不同溶剂中的溶解度选择合适的溶剂配制储备液；一旦配成溶液，请分装保存，避免反复冻融造成的产品失效。
储备液的保存方式和期限：-80°C, 6 months; -20°C, 1 month (protect from light)。-80°C 储存时，请在 6 个月内使用，-20°C 储存时，请在 1 个月内使用。

请根据您的实验动物和给药方式选择适当的溶解方案。以下溶解方案都请先按照 In Vitro 方式配制澄清的储备液，再依次添加助溶剂：

——为保证实验结果的可靠性，澄清的储备液可以根据储存条件，适当保存；体内实验的工作液，建议您现用现配，当天使用； 以下溶剂前显示的百
分比是指该溶剂在您配制终溶液中的体积占比；如在配制过程中出现沉淀、析出现象，可以通过加热和/或超声的方式助溶

• 1.

  请依序添加每种溶剂：;10% DMSO ;; 40% PEG300 ;; 5% Tween-80 ;; 45% saline

  Solubility: ≥ 2.5 mg/mL (13.65 mM); Clear solution

  此方案可获得 ≥ 2.5 mg/mL (13.65 mM，饱和度未知) 的澄清溶液。

  以 1 mL 工作液为例，取 100 μL 25.0 mg/mL 的澄清 DMSO 储备液加到 400 μL PEG300 中，混合均匀；向上述体系中加入50 μL Tween-80，混合均匀；然后继续加入 450 μL生理盐水定容至 1 mL。

  将 0.9 g 氯化钠，完全溶解于 100 mL ddH₂O 中，得到澄清透明的生理盐水溶液

• [1]. Wu X, et al. Determination of amino acid neurotransmitters in rat hippocampi by HPLC-UV using NBD-F as a derivative. Biomed Chromatogr. 2014 Apr;28(4):459-62.

  [2]. Ishikawa T, et al. Development of an LC-MS/MS method for the analysis of free sphingoid bases using 4-fluoro-7-nitrobenzofurazan (NBD-F). Lipids. 2014 Mar;49(3):295-304.

An accurately weighed quantity (0.0183 g) of NBD-F is transferred into a 1 mL centrifuge tube, dissolved in acetonitrile and made up to volume with the same solvent to produce stock solutions of 0.1 M. The solution is protected from light and stored at -20°C until analyzed. A 100 μL aliquot of mixed amino acids solution or sample supernatant, 175 μL of borate buffer solution, 200 μL of acetonitrile and 25 μL of NBD-F working solution are mixed in a 1.5 mL centrifuge tube. The well-mixed solution is allowed to react at 60°C in the water bath for 7 min, excluding light. NBD-F reacts with amino group and enables amino acids to be detected with UV detection. After cooling to room temperature, 10 μL of the solution is injected into the equilibrated HPLC system^[1].

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

• [1]. Wu X, et al. Determination of amino acid neurotransmitters in rat hippocampi by HPLC-UV using NBD-F as a derivative. Biomed Chromatogr. 2014 Apr;28(4):459-62.

  [2]. Ishikawa T, et al. Development of an LC-MS/MS method for the analysis of free sphingoid bases using 4-fluoro-7-nitrobenzofurazan (NBD-F). Lipids. 2014 Mar;49(3):295-304.