Aggregated singletons for automated purification (ASAP) is a purification workflow that provides singleton sample purification, registration and delivery to the materials management department as 30 mM dimethyl sulphoxide solutions for biological screening. The singleton samples submitted are aggregated in a mini-array of 10–12 samples and then are analysed using an automated purification process. The steps in the process include pre-quality control (QC), preparative chromatography, solvent evaporation, reformat dilution and duplication, final QC and final registration. The turnaround time from samples received to delivery to materials management is two or three business days. The final QC data are uploaded to a database and are available to chemists via electronic laboratory notebook software. The final purity, weight recovery and registration information is available in a research database and an e-mail notification of completion is sent to the chemist. ASAP enables a high purification success rate; increases the likelihood of running mini-arrays that generate 5–10 analogues in a final step rather than one or two with the same 2–3 day turnaround time; and provides expert-level service and technology.
The turnaround time from samples received for purification to delivery to the materials management department is two to three business days. Centralizing this activity in the purification group allows greater time for higher value tasks to be completed by practicing chemists; the purification scientists can provide expert-level service and technology, and also more opportunity for harmonization with screening (consistent and high quality samples delivered for biological assays).Experimental
The dried, purified material is dissolved in DMSO to make 30 mM solutions. A maximum of 900 µL of the 30 mM DMSO solution is transferred to a plate. A daughter analytical plate is also created by transferring a 5 µL aliquot and adding enough DMSO to result in a 0.5 mg/mL concentration. Excess 30 mM DMSO solution is transferred to bar-coded 2 mL vials. Tecan liquid handling systems are used for dispensing the DMSO and transfer of samples to the plates and vials. The samples in the vials are evaporated using a Genevac HT-12 evaporator in a two-step evaporation process.
The final QC is performed using the HPLC–MS–ELSD system or Waters Acquity UPLC/SQD/PL 2100 ELSD units, and the samples are registered if they meet the purity criteria (>80% purity by UV at 215 nm, >85% purity by ELSD and >50% mass spectral purity). Orthogonal QC methods are selected to ensure the final purity of the samples. The solubilized plate (maximum up to 900 µL of 30 mM DMSO stock) is sent to the materials management department for assay preparations. The compound is then released for the requested screening. The dry compounds are registered and shipped to the Pfizer Neat Store. The pre-QC and final QC data are uploaded in Pfizer's Global Analytical database as a PDF and the file also gets linked to the chemist's e-Notebook submission page. The recovery amount, purity and registration data are available through Pfizer's Research database. An e-mail notification is sent to the chemists to inform them that the samples have been delivered to the materials management department. A results spreadsheet is attached that provides detailed information regarding the samples, including the QC gradient conditions, mass observed, retention time, purity data and total recovery after purification. The entire process and data flow are handled through customized software. The software has been specifically designed for automated batch-singleton and library purification workflow. The software enables a user to handle multiple plates with accurate data flow. It also provides structures, chemical properties, CLogD data and acid–base labile prediction information for the compounds associated in the plate. This information helps minimize the time required for method development.