Polymer Cookery: Influence of Polymerization Time and Different Initiation Conditions on Performance of Molecularly Imprinted Polymers Sergey A. Piletsky,* Irene Mijangos, Antonio Guerreiro, Elena V. Piletska, Iva Chianella, Kal Karim, and Anthony P. F. Turner Institute of BioScience and Technology, Cranfield University, Silsoe, Bedfordshire, MK45 4DT, U.K. Received September 24, 2004; Revised Manuscript Received November 9, 2004 ABSTRACT: A set of polymers was imprinted with (-)-ephedrine using thermal and UV initiation. The performance of the synthesized materials was studied by HPLC. It was shown that the polymer morphology and enantioselective properties are dependent on the polymerization conditions and time of the reaction. The binding mechanism of synthesized polymers was studied using Van’t Hoff analysis. The results of testing strongly indicate that the polymer receptor structure is shaped during the initial phase of polymer gelation. The present study shows the importance of controlling experimental conditions in the MIP synthesis and highlights potential problems expected during scaling up of MIP production. Introduction During the past 10 years, substantial progress has been made in understanding the nature of the complex phenomena involved in synthesis and related perfor- mance of molecularly imprinted polymers. 1,2 The factors studied include analysis of the type and concentration of the monomers used in polymer preparation, 3,4 role of polymerization temperature, 5,6 pressure 7,8 and sol- vent. 9,10 Surprisingly, little is known about the optimal time needed for MIP formation and its relation to the kinetics of polymerization reaction. This could be explained partially by the difficulty in studying the complex processes involved in the formation of highly cross- linked polymers and the monomer-template complex- ation during polymerization reaction. We have shown before that small differences in the temperature profile of the initial period of the polym- erization reaction causes a profound effect on the polymer morphology and recognition properties. 6 The question which required further attention was the influence of the polymerization time on MIP recognition abilities. It is expected that, irrespective of the initiation protocol (UV or thermopolymerization), the prolonged polymerization time should lead to higher completion of the polymerization reaction with fewer unpolymer- izable double bonds remaining in the polymer. Polymers formed during longer periods of time would be more rigid. This should ensure a more defined shape of imprinting cavities and higher specificity of MIPs. On the other hand, more-rigid polymers might have hin- dered mass exchange and slow binding kinetics. The particular balance of rigidity of imprinting cavities versus polymer flexibility and adaptability (an analogue of induced fit in natural receptors and enzymes) remains poorly understood. The current study is aimed at the analysis of the role of time of polymerization on MIP recognition abilities. The model system includes a set of hydroxyethyl meth- acrylate (HEMA)-based polymers, which have been imprinted with (-)-ephedrine using two different initia- tion protocols: thermal and UV polymerization. The synthesized materials were studied using HPLC, and their morphology, swelling data, and performance were compared. Results and Discussion To analyze the effect of polymerization time on the MIP’s affinity and specificity, three sets of polymers were imprinted with (-)-ephedrine using different initiation protocols: thermoinitiation at 80 °C, photo- initiation with a “strong” UV lamp (0.163 W/cm 2 ), and photoinitiation with a “weak” UV lamp (0.016 W/cm 2 ) at 0 °C. Each set was prepared using different exposure times, which varied from 20 min to 7 days. The polymer’s performance was analyzed using chromato- graphic experiments. The temperature of the initial period of polymeriza- tion reaction was monitored using a thermocouple as described previously (Figure 1). 6 As expected, the real polymerization temperature was significantly higher than the one used for the initiation of the reaction due to the exothermic nature of the process. After the initial 20 min, the reaction temperature monotonically declined with the exception of the “strong” UV lamp initiation protocol, which generated so much heat that the tem- perature continued to increase further up to 200 °C over the next 3 h. This increase did not result from the reaction itself since the measurement of the tempera- ture of mineral oil exposed to the same conditions gave the same temperature increase (190 °C). The photodecomposition of azo compounds could be substantially faster than their thermodecomposition at 80 °C, the condition traditionally used for MIP prepara- tion. Evidence to support this hypothesis comes from a comparison of previously published data describing the behavior of two similar initiators in acetonitrile and toluene. Thus, for a photoinitiation with a medium- pressure Hg lamp at 25 °C, the half-life (t 1/2 ) of azo-bis- (cianovaleric acid) is approximately 140 min. 11 At 80 °C, the thermodecomposition of similar compound, 1,1′- azobis (cyclohexanecarbonitrile) is more than 10 times * Author to whom correspondence should be addressed. Tel: +44 (0)1525 863584. Fax: +44 (0)1525 863533. E-mail: S.Piletsky@cranfield.ac.uk. 1410 Macromolecules 2005, 38, 1410-1414 10.1021/ma048021r CCC: $30.25 © 2005 American Chemical Society Published on Web 01/21/2005