Comparison of Kinetics of Organic Reactions Carried out on Resin

The effect of reagent penetration into the resin on the reaction kinetics was studied by examining four solid-phase organic reactions on resin beads o...
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Ind. Eng. Chem. Res. 2003, 42, 5964-5967

Comparison of Kinetics of Organic Reactions Carried out on Resin Beads of Different Diameters Bing Yan*,† and Qing Tang‡ ChemRx Division, Discovery Partners International, Inc., 385 Oyster Point Boulevard, South San Francisco, California 94080, and Vertex Pharmaceutical Inc., 130 Waverely Street, Cambridge, Massachusetts 02139

The effect of reagent penetration into the resin on the reaction kinetics was studied by examining four solid-phase organic reactions on resin beads of various sizes. In three esterification reactions, the declining trend is observed only for the smallest acid used. As the acids become bulkier, no dependence of the reaction rate on the bead size was observed, indicating that when the reaction rate becomes limiting, the diffusion does not control the kinetics. In a bromination reaction, a reagent salt is composed of a triphenylphosphous cation and a bromide anion. We observed that organic salts are capable of penetrating resin and facilitating the reaction, but there is no clear correlation with bead size. Introduction Combinatorial chemistry1,2 has been accepted as an essential discovery tool. Solid-phase organic synthesis has become a major methodology for synthesizing smallmolecule combinatorial libraries.1,2 However, immense effort is expected to optimize synthesis protocol in order to make a high-quality compound library. Compound library synthesis without going through careful optimization procedures tends to give lower yields and higher impurities in the final products. The low-quality libraries will lead to ambiguity and uncertainty in the biological assay. Reaction kinetics on resin determines the time required for solid-phase synthesis and the likelihood of the reaction completion that determines the final purity and yield of the product. Such information is important for solid-phase combinatorial synthesis using the parallel format as well as using the split-and-pool3-5 method. Recently, the miniaturized screening format allows the use of much less compound for each assay.6,7 Split synthesis using beads with a diameter of 500 µm can provide enough compound to prepare a stock solution for ∼7000 assays. This established that the split synthesis be a more economical method for synthesizing combinatorial libraries for biological screening. However, the reaction rate on such macrobeads may be different from that on regular beads with a diameter of 50-100 µm. In this paper, we report a kinetics study using a single-bead Fourier transform infrared (FTIR) method8-10 on resin beads with various diameters (Figure 1) in order to gain a better understanding of the organic reactions carried out on macrobeads. Results Esterification Reaction on Resin Beads of Various Sizes. In this experiment, cyclopentylacetic acid, 2-norbornaneacetic acid, and 1-admantaneacetic acid * To whom correspondence should be addressed. † Discovery Partners International, Inc. ‡ Vertex Pharmaceutical Inc.

Figure 1. Photographs of resin beads used in this work. Pictures were taken at the same scale. The bead diameters are (1) 35-75 µm, (2) 160-200 µm, and (3) 500-560 µm.

Scheme 1

reacted, respectively, with the alcohol resins 1 (Scheme 1) of different sizes. These resins have different diameter ranges, namely, 35-75, 160-200, and 500-560 µm. The relative conversion of the starting material to the product was determined by a single-bead FTIR method. Figure 2 shows IR spectra taken at various times during the reaction. In all cases, the IR band at ∼3500 cm-1 that represents the starting alcohol on resin decreases its intensity with time. At the same time, the IR band at 1734 cm-1 increases its intensity with time. The reaction process was monitored by integrating IR peak areas for the starting material and the product at various times. The results are shown in Figure 3. Rate constants were obtained by curve-fit analysis of all reactions based on a pseudo-first-order reaction model (Table 1).11 The relationship between the reaction rate and the bead size for three acids in an esterification reaction is shown in Figure 4. When the bulkier acid 7 was used, the reaction was slowed by 30%. Although in

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Figure 2. Single-bead FTIR spectra of resins at various times during the reaction between 1 and 5. Panel A is for the reaction carried out on resins with a diameter of 35-75 µm, panel B is for resins with a diameter of 160-200 µm, and panel C is for resins with a diameter of 500-560 µm. Arrows point out the IR bands that change their intensities during the functional group transformation.

Figure 3. Time courses of all nine esterification reactions and their pseudo-first-order best fits (lines). In each panel, both the decreasing starting resin (solid symbols) and the formation of the product (open symbols) are shown. Panels A, D, and G are for the reaction carried out on resins with a diameter of 35-75 µm, panels B, E, and H are for resins with a diameter of 160-200 µm, and panels C, F, and I are for resins with a diameter of 500-560 µm.

Figure 4. Relationship between the average bead diameter and the reaction rate constant when acids 5-7 were used in the reaction (Scheme 1).

Scheme 2

Table 1. Rate Constants × 10-3 (1/s)a acid 1 bead 1 bead 2 bead 3 a

acid 2

acid 3

bromination

sm

prod

sm

prod

sm

prod

sm

0.99 0.84 0.55

0.88 0.79 0.53

0.60 0.93 0.55

0.53 0.81 0.60

0.41 0.44 0.49

0.38 0.48 0.44

0.57 2.06 1.1

sm ) starting material.

the case of acid 5 there was a slightly reduced reaction rate for larger beads, in the cases of 6 and 7, the bead size was not rate limiting, suggesting that the reagent diffusion was not limiting the reaction. The data, however, did not exclude the possibility that the diffusion rate might become insignificant because the chemical reaction rate was too slow. Bromination Reaction Using Resin Beads of Various Sizes. The bromination reaction was carried out on resins 1 with three different sizes (Scheme 2). Single-bead FTIR spectra were recorded (Figure 5) on

resin beads taken out of the reaction solution at various times during the reaction. The area of the IR band at ∼3500 cm-1 was integrated as the indication for the amount of remaining starting material. The product was further confirmed and quantified by bromine analysis on resin beads.12 Rate constants on various beads were obtained by kinetics analysis as described above (Figure 6 and Table 1). The relationship between the rate constants and the bead size is shown in Figure 7. The size of the resin bead, therefore the diffusion rate, did not limit the reaction rate in this case. Discussion Different chemicals have different tendencies to remain in the bulk or inside the resin. In a solid-phase

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Figure 5. Single-bead FTIR spectra of resins at various times during the reaction between 1 and 8. Panel A is for reaction carried out on resins with a diameter of 35-75 µm, panel B is for resins with a diameter of 160-200 µm, and panel C is for resins with a diameter of 500-560 µm. Arrows point out the IR bands that change their intensities during the functional group transformation.

Figure 7. Relationship between the average bead diameter and the reaction rate constant for the reaction shown in Scheme 2.

Figure 6. Time courses of all three bromination reactions and their pseudo-first-order best fits (lines). In each panel, the relative amount of the starting resin (solid symbols) is shown. Panels A-C are for reactions carried out on resins with diameters of 35-75, 160-200, and 500-560 µm, respectively.

organic reaction, reagent molecules need to (1) have a high tendency to be absorbed into the resin, (2) diffuse to the reaction site, and (3) react with the resin-bound reactant. Besides the intrinsic reactivity, the rate and efficiency of absorption and diffusion processes will affect the reaction kinetics. The higher local concentration of reagents in the resin will give a higher reaction rate.13 Factors that determine the local concentration inside the resin are not well-known. Charge carried by the resin or reagent may affect the absorption. If the gel phase of swollen resin can be considered as another

solvent phase,14 the principle of “like dissolve like” will be true for resin absorption selectivity. A polar compound had a higher absorptivity in polar resins.15 Experimental data have shown that solvent molecules diffused inside the resin bead at a different self-diffusion coefficient compared to that outside the resin.16 The diffusion of reagent molecules may also differ. Theoretically, reagent diffusion should vary depending on the bead size. This was experimentally demonstrated using an acylation reaction.17 Molecular diffusion also depends on the extent of resin swelling.18 Resin swelling depends on the base resin [polystyrene (PS)-2% DVB in here] and the attached functional group. In this study, the base resin and the functional group are identical. The loading of the functional group is similar. Even though the sizes of these resins differ, the relative swelling can be assumed to be close. In the esterification experiment, the reaction rate is the same for resin beads of different sizes when a specific acid was used. The kinetics only reflects the absorption rate and the diffusion rate. The difference in loading for resins with different diameters was corrected by normalizing loading for all resins. The declining trend is observed for the smallest acid used in the esterification reaction. As the acids become bulkier, the reaction rate decreases. No dependence of the reaction rate on the bead size is observed. This indicates that when the reaction rate becomes limiting, the diffusion does not control the kinetics. In the bromination reaction, the

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reactive intermediate is a bromide salt formed between triphenylphosphine and carbon tetrabromide.19 However, the salt is composed of a triphenylphosphous cation and a bromide anion. Both species are “soft”. We have observed that some “hard” salts species such as sodium acetate have difficulty penetrating into the resin. Here we see that organic salts are capable of penetrating the resin and facilitating the reaction. In summary, four organic reactions were carried out on resin beads of various sizes. In three of four cases, the reaction rate is similar and the diffusion is not limiting the reaction.

diamond window (SpectraTech, Shelton, CT) was used to collect the background spectrum. Data were collected at 4 cm-1 resolution, and 32 scans were averaged for each bead. (a) Data Analysis. The ratio of an IR band peak area to the peak area of a PS band at 1947 cm-1 was used as a measure for the amount of compound. This ratio was used to quantify the percentage conversion. Average integrated peak areas from three to five beads were normally used to plot against time for kinetic analysis. These data points were fitted to a pseudo-first-order rate equation by using SigmaPlot (Jandel Scientific, San Rafael, CA) on a PC.

Experimental Section Materials. Rapp A-OH resins were purchased from RAPP Polymere Gmbh. These resins were based on cross-linked PS with 1% DVB. Their diameters and loadings are as follows: bead 1, 35-75 µm, 1.48 mmol/ g; bead 2, 160-200 µm, 1.26 mmol/g; bead 3, 500-560 µm, 1.14 mmol/g. Other chemical reagents were purchased from Aldrich and used as received. Resins were preswollen in dimethylformamide (DMF) or tetrahydrofuran (THF) for 30 min before reactions. (a) Esterification Reaction. The resin (0.057 mmol) was suspended in DMF (2 mL). To the suspension was added the acid (0.57 mmol), DMAP (70 mg, 0.57 mmol), and DIC (0.09 mL, 0.57 mmol). The resulting suspension was rotated on a Glas-Col laboratory rotor shaker (32 rpm). A small amount of resin was removed at 5, 10, 30, 1.5, and 4 h. The resin sample was washed with DMF (5 × 0.5 mL), THF (5 × 0.5 mL), and CH2Cl2 (10 × 0.5 mL) and used for single-bead FTIR analysis. (b) Bromination Reaction. To a solution of CBr4 (114 mg, 0.34 mmol) in THF (1 mL) was added a solution of PPh3 (90 mg, 0.34 mmol) in THF (1 mL). The resulting mixture was poured into the resins (0.114 mmol) immediately. The resin suspension was rotated on a Glas-Col laboratory rotor shaker (32 rpm). A small amount of resin was removed at 5, 10, 30, 1.5, and 4 h. The resin sample was washed with DMF (5 × 0.5 mL), THF (5 × 0.5 mL), and CH2Cl2 (10 × 0.5 mL) and used for single-bead FTIR analysis (Scheme 2). Single-Bead FTIR Method. FTIR spectra were collected on a Nicolet Nexus 670 with a continuum microscope, using OMNIC software. The microscope is equipped with a 15× cassegrain objective and a liquidnitrogen-cooled mercury-cadmium-telluride detector. The view mode aided in locating a single bead. The transmission mode was used for the whole bead measurement. Beads were flattened with a diamond window (SpectraTech, Shelton, CT). A clean area of the same

Acknowledgment We thank Ms. Lina Liu for her technical assistance. Literature Cited (1) Dolle, R. J. Comb. Chem. 2001, 3, 477. (2) Dolle, R. J. Comb. Chem. 2000, 2, 383. (3) Furka, A.; Sebestyen, F.; Asgedom, M.; Dibo, G. Int. J. Pept. Protein Res. 1991, 37, 487. (4) Houghten, R. A.; Pinilla, C.; Blondelle, S. E.; Appel, J. R.; Dooley, C. T.; Cuervo, J. H. Nature 1991, 354, 84. (5) Lam, K. S.; Salmon, S. E.; Hersh, E. M.; Hruby, V. J.; Kazmierski, W. M.; Knapp, R. J. Nature 1991, 354, 82. (6) Hergenrother, P. J.; Depew, K. M.; Schreiber, S. L. J. Am. Chem. Soc. 2000, 122, 7849-7850. (7) MacBeath, G.; Koehler, A. N.; Schreiber, S. L. J. Am. Chem. Soc. 1999, 121, 7967-7968. (8) Yan, B.; Kumaravel, G.; Anjaria, H.; Wu, A.; Petter, R.; Jewell, C. F., Jr.; Wareing, J. R. J. Org. Chem. 1995, 60, 5736. (9) Yan, B. Acct. Chem. Res. 1998, 31, 621. (10) Yan, B.; Kumaravel, G. Tetrahedron 1996, 52, 843. (11) Yan, B.; Fell, J. B.; Kumaravel, G. J. Org. Chem. 1996, 61, 7467. (12) Yan, B.; Jewell, C. F., Jr.; Myers, S. W. Tetrahedron 1998, 54, 11755. (13) Cainelli, G.; Contento, M.; Manoscalchi, F.; Plessi, L. J. Chem. Soc., Chem. Commun. 1982, 725. (14) Czarnik, A. W. Biotechnol. Bioeng. 1998, 61, 77. (15) Regen, S. L.; Nigam, A. J. Am. Chem. Soc. 1978, 100, 7773. (16) Ford, W. T.; Ackerson, B. J.; Blum, F. D.; Periyasamy, M.; Pickup, S. J. Am. Chem. Soc. 1987, 109, 7276. (17) Wilson, M. E.; Paech, K.; Zhou, W.-J.; Kurth, M. J. J. Org. Chem. 1998, 63, 5094. (18) Groth, T.; Grotli, M.; Meldal, M. J. Comb. Chem. 2001, 3, 461. (19) Ramirez, F.; Desai, N. B.; McKelvie, N. J. Am. Chem. Soc. 1962, 84, 1745.

Received for review February 18, 2003 Revised manuscript received May 21, 2003 Accepted July 27, 2003 IE030149U