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Communications Synthesis of Poly(2-methyl-3-hydroxyoctanoate) via Anionic Polymerization of r-Methyl-β-pentyl-β-propiolactone Ali Hakan Arkin,†,‡ Baki Hazer,‡,§ Graz˘ yna Adamus,| Marek Kowalczuk,| Zbigniew Jedlin´ ski,*,| and Robert W. Lenz⊥ Department of Chemistry, Zonguldak Karaelmas University, 67100 Zonguldak, Turkey; Food Science and Technologies Research Institute, TUBITAK-Marmara Research Center, Gebze 41470 Kocaeli, Turkey; Centre of Polymer Chemistry, Polish Academy of Sciences, 41-800 Zabrze, Poland; and Polymer Science and Engineering Department, University of Massachusetts, Amherst, Massachusetts 01003-4530 Received February 28, 2001; Revised Manuscript Received June 18, 2001
Synthesis of an R,β-alkyl branched polyester, i.e., poly(2-methyl-3-hydroxyoctanoate), has been accomplished via anionic polymerization of R-methyl-β-pentyl-β-propiolactone mediated by supramolecular complexes of potassium methoxide or potassium hydroxide, respectively. The structure of resulting polymers has been established by electrospray ionization multistage mass spectrometry (ESI-MSn), FT-IR, NMR, and GPC analyses. Previously proposed addition-elimination mechanism of the polymerization of β-lactones containing R-hydrogen by alkoxide anion has been confirmed to operate also in the case of β-lactone having alkyl substituents in both R and β positions. Natural poly [(R)-3-hydroxyalkanoates], PHA, are widely distributed in biological systems, high molecular weight PHA being produced by a wide range of microorganisms as intracellular carbon and energy sources.1,2 The attempts to incorporate various functional groups into the structure of PHA in order to improve their properties were undertaken during the past few decades.3-7 Structural modification of PHA via R-methyl branching was accomplished previously by one of us, and the copolymer containing up to 5% of 2-methyl-3-hydroxyoctanoate structural units was prepared with the aid of co-feeding technique of Pseudomonas oleoVarans.7 It was also found that R-methyl-branched PHA copolymers exhibit different crystallinity and melting transitions than those of respective non-R-substituted PHA. The ring-opening polymerization (ROP) of respective β-lactones constitutes an alternative approach to PHA synthesis with respect to biofermentation.8 Chemical synthesis of functionalized poly [(R,S)-3-hydroxybutyrate], a-PHB, and its copolymers via ring-opening of β-butyrolactone (ROP) mediated by activated anionic initiators was demonstrated.9 Using this synthetic approach, a-PHB with defined chemical structure as well as block, graft, and random copolymers have been obtained and characterized10 Recently * Corresponding author. Telephone: +48322716077. Fax: +48322712969. E-mail:
[email protected]. † On temporary leave to the Centre of Polymer Chemistry, Polish Academy of Sciences, under the framework of TUBITAK NATO-A2 research grant. ‡ Zonguldak Karaelmas University. § TUBITAK-Marmara Research Center. | Polish Academy of Sciences. ⊥ University of Massachusetts.
Scheme 1
a novel facile method of regioselective synthesis of stereoregular biomimetic [R]-PHB, analogous to the natural one, has also been demonstrated.11 In the present work we report on the synthesis and structural studies of the analogue of natural R-methylbranched PHA, i.e., poly(2-methyl-3-hydroxyoctanoate), 2, obtained for the first time to our knowledge via anionic polymerization of racemic R-methyl-β-pentyl-β-propiolactone (1).12 Supramolecular complexes of potassium methoxide and potassium hydroxide with 18-crown-6 have been selected as respective activated anionic initiators of the polymerization (Scheme 1).13 On the basis of polymer end groups analysis, the polymerization mechanism has been compared with that proposed previously for ROP of simple β-lactones, i.e., β-propiolactone and β-butyrolactone, conducted in the presence of these anionic initiators.14-16 Two samples of poly(2-methyl-3-hydroxyoctanoate) 2, obtained via anionic polymerization of the racemic lactone 1 by these initiators were selected for structural studies. Sample 1, obtained with the aid of a supramolecular complex
10.1021/bm015528q CCC: $20.00 © 2001 American Chemical Society Published on Web 07/06/2001
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Figure 1. (a) ESI-MS spectrum (negative ion-mode) of polymer 2 prepared in the polymerization of lactone 1 initiated by KOH/18-crown-6 complex. (b) Expanded ESI-MS spectrum of polymer 2 prepared in the polymerization of lactone 1 initiated by CH3OK/18-crown-6 complex. (c) Expanded ESI-MS spectrum of polymer 2 prepared in the polymerization of lactone 1 initiated by KOH/18-crown-6 complex.
18-crown-6 with potassium methoxide possessed the molecular weight Mn ) 2100 and the polydispersity index Mw/Mn ) 1.2. Sample 2, obtained in the presence of a supramolecular complex 18-crown-6 with potassium hydroxide, had Mn ) 1200 and Mw/Mn ) 1.2, as revealed by the GPC experiments. Polymers obtained were characterized by FT-IR, 1H NMR, and 13C NMR.13 However, the characterization of the end groups in the polymer 2 by NMR method was difficult, due to the overlapping of the respective resonance signals, and no detailed information was provided. Therefore, electrospray ion trap multistage mass spectrometry (ESI-MSn) was applied for the structural studies of the polymer 2. The usefulness of this technique for structural studies of mass-selected macromolecules of chemically synthesized as well as natural aliphatic polyesters has been recently demonstrated.17-21 Using the ESI-MSn technique, determination of molecular masses and structures of massselected macromolecular ions of polyester 2 has been accomplished, thus showing the chemical nature of the polymer and its end groups. The ESI-MS analysis (performed in negative ion mode) revealed that despite of the anionic initiator used (CH3OK/ 18-crown-6 for sample 1 and KOH/18-crown-6 for sample 2) the poly(2-methyl-3-hydroxyoctanoate) 2 contained two kinds of macromolecular chains (Scheme 1, Figure 1,
macromolecules 2a and 2b) with different end groups, i.e., 2-methyl-3-hydroxyoctanoate (Scheme 1 and Figure 1, 2a) and unsaturated 2-methyl-2-octenoate (Scheme 1 and Figure 1, 2b), respectively.22 The presence of two sets of anions corresponding to two kinds of individual macromolecular chains (2a and 2b) of the general chemical structure presented in Figure 1 were observed. The peaks corresponding to each set showed a peak-to-peak mass increment of 156 Da, which is equal to the molecular weight of the 2-methyl-3-hydroxyoctanoate repeat unit. The mass difference between the m/z values of neighboring anions of individual macromolecular chains of 2a and 2b, having the same degree of polymerization, equals 18 Da which is the same as the difference between the molecular weight of 2-methyl-3-hydroxyoctanoate and 2-methyl-2-octenoate end groups (Figure 1a). No molecular ions of the macromolecules of polymer 2 contained incorporated methoxide initiator were observed in the ESI-MS spectrum of sample 1 (Figure 1b), and despite the anionic initiator used, the MS spectra of the polymers obtained (samples 1 and 2) were almost identical (compare parts b and c of Figure 1). The correctness of the above assignment was provided by the multistage ESI-MSn experiments performed for two selected parent anions at m/z 797 and m/z 779 corresponding
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Figure 2. Sequential fragmentation spectra MSn (negative ion-mode) obtained for selected parent ion m/z 797 of polymer 2a. Scheme 2
to the set 2a and 2b, respectively (Figure 1). The pertinent sequential (MSn) spectra are presented in Figures 2 and 3,
respectively, and the proposed fragmentation mechanism is depicted in Scheme 2.
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polymerization of β-lactone 1 initiated by selected activated anionic initiators. This synthetic approach constitutes an alternative way to respective R-methyl-branched polyesters as compared with biofermentation by P. oleoVarans. On the basis of the presented results previous assumptions and views on the chemistry of β-lactones ring-opening polymerization have been corrected.23
Figure 3. Sequential fragmentation spectrum MS2 (negative ionmode) obtained for selected parent ion m/z 779 of polymer 2b.
The presence of 2-methyl-3-hydroxyoctanoate end group in polymer 2a was confirmed by multistage MS experiments performed for selected parent anion at m/z 797 (Figure 2). Because of the fragmentation at both ends of the 2a polymer molecule (Scheme 2 and Figure 2: MS2) the fragment anion at m/z 641 is formed as a result of the loss of 2-methyl-2-octenoic acid (156 Da), and the fragment anion m/z 623 indicates the expulsion of 2-methyl-3-hydroxyoctanoic acid (174 Da). Further fragmentation of the anion m/z 641 (containing 2-methyl-3-hydroxyoctanoate and carboxylate end groups) creates again two sets of fragment anions (Scheme 2 and Figure 2: MS3). However, further fragmentation of the fragment anion m/z 623 (containing 2-methyl-2-octenoate and carboxylate end groups) creates only one set of fragment anions due to successive losses of 2-methyl-2-octenoic acid from both sides of polymer molecule (Scheme 2 and Figure 2: MS3 of m/z 623). The ESI-MS2 fragmentation experiment conducted for the selected parent anion of polymer 2b contained 2-methyl2-octenoate and carboxylate end groups (Figure 1, m/z 779) indicated only one set of fragment anions at m/z 623, 467, and 311 formed by successive losses of 2-methyl-2-octenoic acid (Figure 3, MS2). The results of ESI-MSn experiments presented above demonstrated that regardless of the anionic initiator used (18crown-6 supramolecular complex of potassium hydroxide or potassium methoxide), poly(2-methyl-3-hydroxyoctanoate) 2, prepared via anionic polymerization of the lactone 1, contains 2-methyl-3-hydroxyoctanoate and unsaturated 2-methyl-2-octenoate end groups. Thus, it was demonstrated that the mechanism of anionic polymerization of R-methylsubstituted β-propiolactone 1 in the presence of the employed alkali metal alkoxide is similar to that previously proposed by some of us for simple β-lactones,14-16 and potassium hydroxide acts as a real initiator in this system. The results presented in this communication constitute further extension of the previously proposed addition-elimination mechanism of the polymerization of β-lactones containing R-hydrogen and carried out in the presence of alkoxide anion, since clearly macromolecules with either hydroxyl or unsaturated end groups are formed. Conclusions The results of the present study revealed that it is possible to obtain the polymer 2 via the regioselective ring-opening
Acknowledgment. Financial support from the EUREKA E! 2004 “MICROPOL” Grant and U.S.-Polish M. Skłodowska-Curie Joint Fund II, Grant PAN-NSF No. 94/195 is acknowledged. References and Notes (1) (2) (3) (4) (5) (6)
(7) (8) (9) (10) (11) (12)
(13)
(14) (15)
Byrom, D. Trends Biotechnol. 1987, 5, 246. Sudes, K.; Abe, H.; Doi, Y. Prog. Polym. Sci. 2000, 25, 1503. Hazer, B.; Lenz, R. W.; Fuller, R. C. Polymer 1996, 37, 5951. Curley, J. M.; Hazer, B.; Lenz, R. W.; Fuller, R. C. Macromolecules 1996, 29, 1762. Takagi, Y.; Hashii, M.; Maehara, A.; Yamane, T. Macromolecules 1999, 32, 8315 and references cited therein. (a) Hazer, B.; Lenz, R. W.; Fuller, R. C. Macromolecules 1994, 27, 45. (b) Fritzsch, K.; Lenz, R. W.; Fuller, R. C. Int. J. Biol. Macromol. 1990, 12, 92. (c) Ibaoglu, K.; Hazer, B.; Arkin, A. H.; Lenz, R. W. Bull. Chem. Technol. Maced. 2000, 19, 41. Scholz, C.; Wolk, S.; Lenz, R. W.; Fuller, R. C. Macromolecules 1996, 27, 6358. (a) Lenz, R. W.; Jedlin´ski, Z. Macromol. Symp. 1996, 107, 149. (b) Peres, R.; Lenz, R. W. Polymer 1994, 35, 1059. Jedlin´ski, Z. Acta Chem. Scand. 1999, 53, 157. Jedlin´ski, Z.; Kowalczuk, M.; Adamus, G.; Sikorska, W.; Rydz, J. Int. J. Biol. Macromol. 1999, 25, 247. Jedlin´ski, Z.; Kurcok, P.; Lenz, R. W. Macromolecules 1998, 31, 6718. The R-methyl-β-pentyl-β-propiolactone (3-methyl-4-pentyl-2-oxetanone), 1, was obtained from 2-methyl-3-hydroxyoctanoic acid according to the literature (Adam, W.; Baeza, J.; Liu, Ju-C. J. Am. Chem. Soc. 1972, 94, 2000). Bp: 66 °C (0.3 mmHg). The spectral data of lactone 1 were as follows: IR (cm-1, neat) 1827 (s, carbonyl), 1149 (m, C-O). 1H NMR (300 MHz) δ (CDCl3, TMS, ppm), 0.88 (t, 3H, terminal CH3 of the pentyl group, J ) 6.84 Hz), 1.24-1.26 and 1.35-1.37 (dd, 3H, R-CH3, J ) 10.6 Hz), 1.25-1.52 (m, 6H, internal CH2 of the pentyl group), 1.58-1.86 (m., 2H, γ-CH2), two sets of multiplets between 3.15 and 3.25 and 3.67-3.78 (1H, R-H), and two sets of multiplets between 4.12 and 4.19 and 4.49-4.57 (1H, β-H). General polymerization procedure: Poly(2-methyl-3-hydroxyoctanoic acid), 2, was synthesized by anionic ring-opening polymerization of 3-methyl-4-pentyl-β-propiolactone, 1, with KOH/18-crown-6 or CH3OK/18-crown-6 complexes as initiators, respectively. The polymerization was carried out in bulk at a temperature of 70 °C, and the extent of the reaction was monitored by FT-IR spectroscopy. After completion of the reaction, the polymer was dissolved in chloroform, and the ion-exchange resin in acid form was added as the termination agent and 18-crown-6 adsorber. The polymer obtained was characterized by ESI-MSn, NMR (Varian VCR-300 multinuclear spectrometer), gel permeation chromatography (GPC), and Fourier transform infrared spectroscopy (FT-IR). The spectral data were as follows: FT-IR (cm-1, neat) 1738 (s, carbonyl); 1H NMR (300 MHz) δ (CDCl3, TMS, ppm), 0.87 (t, 3H, terminal CH3 of the pentyl group, J ) 6.28 Hz), 1.13-1.17 (m., 3H, R-CH3), 1.20-1.36 (m., 6H, internal CH2 of the pentyl group), 1.48-1.66 (m., 2H, γ-CH2), 2.582.85 (m, 1H, R-H), 5.03-5.23 (m., 1H, β-H). Number-average molecular mass (Mn) and polydispersity index (Mw/Mn) were estimated by GPC experiments conducted in THF solution at 35 °C, at a flow rate of 1 mL/min using a Spectra-Physics 8800 solvent delivery system with a Plgel 3 µm MIXED-E ultrahigh efficiency column and a Shodex SE 61 refractive index detector. Polystyrene standards with low polydispersities were used to generate a calibration curve. Kurcok, P.; Kowalczuk, M.; Hennek, K.; Jedlin´ski, Z. Macromolecules 1992, 25, 2017. Kurcok, P.; Jedlin´ski, Z.; Kowalczuk, M. J. Org. Chem. 1993, 58, 4219.
Communications (16) Jedlin´ski, Z.; Kowalczuk, M.; Kurcok, P.; Adamus, G.; Matuszowicz, A.; Sikorska, W.; Gross, R. A.; Xu, J.; Lenz, R. W. Macromolecules 1996, 29, 3773. (17) Jedlin´ski, Z.; Adamus, G.; Kowalczuk, M.; Schubert, R.; Szewczuk, Z.; Stefanowicz, P. Rapid Commun. Mass Spectrom. 1998, 12, 357. (18) Arslan, H.; Adamus, G.; Hazer, B.; Kowalczuk, M. Rapid Commun. Mass Spectrom. 1999, 13, 2433. (19) Adamus, G.; Kowalczuk, M. Rapid Commun. Mass Spectrom. 2000, 14, 195. (20) Focarete, M. L.; Scandola, M.; Jendrossek, D.; Adamus, G.; Sikorska, W.; Kowalczuk, M. Macromolecules 1999, 32, 4814. (21) Adamus, G.; Sikorska, W.; Kowalczuk, M.; Montaudo, M.; Scandola, M. Macromolecules 2000, 33, 5797. (22) ESI-MSn experiments: Electrospray mass spectrometric analysis
Biomacromolecules, Vol. 2, No. 3, 2001 627 (ESI-MS) was performed with a Finnigan LCQ ion trap mass spectrometer (Finnigan, San Jose, CA). The polymer sample was dissolved in chloroform methanol system (10:1 v/v) and solution (1.0 mg/mL) was introduced to the ESI source by continuous infusion by means of the instrument syringe pump at a rate of 3 µL/min. The ESI source was operated at 4.25 kV, and the capillary heater was set to 200 °C. For ESI-MSn experiments, mass selected monoisotopic molecular adduct ions were isolated in the ion trap and collisionally activated with 32% ejection RF-amplitude at standard He pressure. The experiments were performed in the negative-ion mode. (23) Hesieh, H. L.; Quirk, R. P. Anionic Polymerization; M. Dekker, Inc.: New York, 1996; p 695.
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