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Fig. 2. Crude synthetic and preparative purified (inset) TAS. Gradient (B = 35 – 65 % in30 min) A = 0.1% TFA, B = 0.1% TFA in 90% aq. ACN; Flow rate: 0.3 mL/min; Max.pressure: 69 kgf/cm 2 .coupling reagents (Aib was coupled with HATU) on TentaGel S RAM in DMF as asolvent. After cleavage the peptide was precipitated from ether to give the crude material(Figure 2). Several test columns (2 i.d. x 250 cm) were prepared and, for comparison,commercial ODS was also used. Based on the above tests a novel reverse phase silica forpreparative separation has been developed and designated Daisogel ® SP (octadecyl silica,10 micrometer particles, 100 and 120 angstrom porosity) (Table 2). The recovery wascalculated by the yield after re-chromatography. Both materials gave >65% recovery andthe 100 angstrom material was slightly better than the 120 angstrom product for the modelpeptide. The homogeneity of the resulting purified peptides was confirmed by LCMS usingthe above HiPep-Cadenza (Figure 2 inset) to give >95% purity (calculated from peak area).These materials were packed in columns (5 i.d. x 50 cm) and several peptides were purifiedon a gram scale with flow rates of 35-50 mL/min.Table 2. Yield from crude material and recovery in the re-chromatography. Freshly packedcolumn was used for preparative purification and the same column was used as the secondseparation, re-chromatography of the purified material.Packing material (10 micron)Yield from crudepeptideRecoveryDaisogel®SP-100-ODS-P (100A) 14.3% 68.2%Daisogel®SP-120-ODS-RPS (120A) 11.8% 66.4%Commercially available 100A-C18(from a leading company)11.1% 32.5%137