Flash chromatography fraction monitoring

An aqueous extract of P. hysterophorus (collected in Puerto Rico) was partitioned into methylene chloride at pH 7, pH 10 and pH 2. Bioassays of the methylene chloride soluble fractions, using the bean second internode bioassay (13), showed that the highest activity was concentrated in the methylene chloride extract at pH 7. Extensive chromatographic purification (flash chromatography, medium pressure LC, preparative TLC) monitored by bioassay led to the isolation of the four sesquiter-... 

The crude product (10 g) is diluted with 4 mL of a solvent mixture (ethyl acetate/cyclohexane = 9 1). This solution is poured onto a column (75-mm diameter) filled with 120 g of silica gel (Merck 230-400 mesh) for flash chromatography. Elution is performed under gravity and requires 200 mL of the above solvent system, followed by 200 mL of ethyl acetate. 2-Phenyl-2-propanol mixed with methyl p-tolyl sulfide is eluted in the first fraction ( 150 mL, monitored by TLC). The subsequent fractions are collected ( 300 mL)... 

When the separation procedures described in detail above are unsatisfactory for the separation of a mixture of organic compounds, purely physical methods may be employed. Thus a mixture of volatile liquids may be fractionally distilled (compare Section 2.26) or a mixture of non-volatile solids may frequently be separated by making use of the differences in solubilities in inert solvents. The progress of such separations may be monitored by application of the various chromatographic techniques detailed in Section 2.31, or indeed these techniques may be employed on the preparative scale for effecting the separation itself (e.g. flash chromatography, p. 217). The techniques of counter current distribution, fractional crystallisation or fractional sublimation (Section 2.21) may also be employed where appropriate. 

For flash chromatography (see Chapter 11), tic is first used to determine the solvent system and quantity of silica required, and secondly to monitor the column fractions. 

The resulting carbamate salts 235 were then reacted with a freshly prepared solution of the activated betaine of TPP or TBP and DIAD to provide the isocyanates 236. After completion of the reaction, as monitored by the IR stretch of the isocyanate, the desired products were obtained by fractional distillation or flash chromatography. Aniline, benzylamine and 2,6-diisopropylaniline gave no or very poor yields of the desired isocyanates under these reaction conditions. The poor yields in these reactions are due to formation of nonreactive intermediate carbamoylhydrazines or competing triazolinone formation the latter arises from reaction of an activated arylisocyanate and the Mitsunobu betaine. 

NaOH (2 ml) and stirred at 60 °C for 10 min after which PMe3 (1 M solution in THF, 3.73 ml, 3.73 mmol) was added. The reaction was monitored hy TLC (DCM/MeOH/HjO/MeNHj (33% solution in EtOH), 10 15 6 15 in EtOH, product J f = 0.33), which indicated completion after 3.5 h. The reaction mixture was then purified by flash chromatography on a short sihca column that was eluted as follows THF, EtOAc, MeOH/EtOAc (1 1), MeOH, and finally with MeNHj (33% solution in EtOH). The fractions containing the product were evaporated under vacuum, redissolved in water, and evaporated again to afford the product in its free amine base form (48.7 mg, 81%). This product was then dissolved in water, the pH was adjusted to 7.5 with 0.01 M H2SO4, and lyophihzed to give the sulfate salt of 99 (88.5 mg) as a white foamy sohd. 

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