Process for the preparation of copolymers derived from 2,4- or 2,5-pyridinedicarboxylic acid
11807886 · 2023-11-07
Assignee
Inventors
Cpc classification
C12Y113/11
CHEMISTRY; METALLURGY
C08G63/6856
CHEMISTRY; METALLURGY
International classification
Abstract
The present invention relates to processes for the formation of pyridinedicarboxylic acid (PDCA), in particular, 2,4-pyridinedicarboxylic acid (2,4-PDCA) and 2,5-pyridinedicarboxylic acid (2,5-PDCA), and mono- and diester derivatives thereof, from 3,4-dihydroxybenzoic acid, via a biocatalytic reaction using, for example, a protocatechuate dioxygenase such as protocatechuate 4,5-dioxygenase or protocatechuate 2,3-dioxygenase, and a nitrogen source. The invention also relates to copolymers that comprise the pyridinedicarboxylic acid monomers and derivatives thereof, processes for the formation of the copolymers and uses for the copolymers.
Claims
1. A method comprising combining: (a) at least one pyridinedicarboxylic acid (PDCA) or a mono- or diester of pyridinedicarboxylic acid; (b) at least one diol; and (c) optionally, at least one aliphatic dicarboxylic acid or a mono- or diester derivative thereof, under conditions suitable for condensation, transesterification or polymerization to produce a copolymer comprising residues of (a), (b) and optionally (c), wherein: the method is catalyzed by an organic acid, an inorganic acid or a metal; the method comprises holding (a), (b) and optionally (c) at a temperature of about 60° C. to about 250° C.; and the method comprises holding (a), (b) and optionally (c) at a pressure of about 1 mbar to about 500 mbar.
2. The method as claimed in claim 1, wherein the copolymer comprises the copolyester of (a) at least one pyridinedicarboxylic acid or a mono- or diester of pyridinedicarboxylic acid; (b) at least one diol; and (c) at least one aliphatic dicarboxylic acid or a mono- or diester derivative thereof.
3. The method as claimed in claim 1, wherein the pyridinedicarboxylic acid or a mono- or diester of pyridinedicarboxylic acid is ##STR00024## wherein each R.sup.1 independently represents H or a straight, branched or cyclic, C.sub.1 to C.sub.6 alkyl group.
4. The method as claimed in claim 1, wherein the pyridinedicarboxylic acid or a mono- or diester of pyridinedicarboxylic acid is selected from the group consisting of ##STR00025## or a combination thereof.
5. The method as claimed in claim 1, wherein the diol is ##STR00026## wherein R.sup.2 is a straight, branched or cyclic C.sub.2 to C.sub.10 alkylene.
6. The method as claimed in claim 1, wherein the diol is selected from the group consisting of 1,2-ethanediol, 1,4-butanediol, and a combination thereof.
7. The method as claimed in claim 1, wherein the aliphatic dicarboxylic acid or a mono- or diester derivative thereof is ##STR00027## wherein R.sup.3 is a straight, branched or cyclic, C.sub.1 saturated or C.sub.2 to C.sub.10 saturated or unsaturated alkylene, and wherein each R.sup.4 independently represents H or a straight, branched or cyclic, C.sub.1 to C.sub.6 alkyl group.
8. The method as claimed in claim 1, wherein the aliphatic dicarboxylic acid or a mono- or diester derivative thereof is selected from the group consisting of adipic acid, adipic acid monomethyl ester, adipic acid dimethyl ester, adipic acid monoethyl ester, adipic acid diethyl ester, succinic acid, succinic acid monomethyl ester, succinic acid dimethyl ester, succinic acid monoethyl ester, succinic acid diethyl ester, and a combination thereof.
9. The method as claimed in claim 1, wherein the copolymer comprises the copolyester of (A) pyridinedicarboxylic acid dimethyl ester or diethyl ester; (B) 1,4-butanediol; and (C) adipic acid dimethyl ester or diethyl ester.
10. The method as claimed in claim 1, wherein the copolymer comprises the copolyester of (A′) pyridinedicarboxylic acid dimethyl ester or diethyl ester; and (B′) 1,4-butanediol or 1,2-ethanediol.
11. The method as claimed in claim 1, wherein the copolymer comprises the copolyester of (a) from 1 to 99 mol % of at least one pyridinedicarboxylic acid or a mono- or diester of pyridinedicarboxylic acid; (b) from 1 to 99 mol % of at least one diol; and (c) when present, from 1 to 98 mol % of at least one aliphatic dicarboxylic acid or a mono- or diester derivative thereof.
12. The method as claimed in claim 1, wherein the pyridinedicarboxylic acid is obtained by a process comprising the steps of (i) contacting a protocatechuate dioxygenase produced by Rhodococcus jostii with 3,4-dihydroxybenzoic acid to form a ring-opened product of 3,4-dihydroxybenzoic acid; and (ii) cyclising the ring-opened product of step (i) with a nitrogen source to form the pyridinedicarboxylic acid.
13. The method as claimed in claim 12, wherein (a) the pyridinedicarboxylic acid is 2,4-pyridinedicarboxylic acid, and wherein the ring-opened product is 4-carboxy-2-hydroxymuconate-6-semialdehyde (CHMS) ##STR00028## or (b) the pyridinedicarboxylic acid is 2,5-pyridinedicarboxylic acid, and wherein the protocatechuate dioxygenase is protocatechuate 2,3-dioxygenase, and the ring-opened product is 5-carboxy-2-hydroxymuconate-6-semialdehyde (5-CHMS) ##STR00029##
14. The method as claimed in claim 1, wherein the mono- or diester of pyridinedicarboxylic acid is obtained by a process comprising the step of (i) providing pyridinedicarboxylic acid; and (ii) adding an alcohol and a catalyst to the pyridinedicarboxylic acid provided in step (i), wherein the pyridinedicarboxylic acid is obtained by (iii) contacting a protocatechuate dioxygenase produced by Rhodococcus jostii with 3,4-dihydroxybenzoic acid to form a ring-opened product of 3,4-dihydroxybenzoic acid; and (iv) cyclising the ring-opened product of step (i) with a nitrogen source to form the pyridinedicarboxylic acid.
15. The method as claimed in claim 1, wherein the method comprises holding (a), (b) and optionally (c) at about 60° C. to about 250° C. for a time period from about 1 hour to about 24 hours.
Description
DESCRIPTION OF THE FIGURES
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(19) The following examples are merely illustrative examples of the invention described herein, and are not intended to be limiting upon the scope of the invention.
(20) Any listing or discussion of an apparently prior-published document in this specification should not necessarily be taken as an acknowledgement that the document is part of the state of the art or common general knowledge. All references disclosed herein are to be considered to be incorporated herein by reference.
Example 1
Synthesis of 2,4-pyridinedicarboxylic acid (2,4-PDCA) and 2,5-pyridinedicarboxylic acid (2,5-PDCA)
(21) ##STR00019##
Introduction of Recombinant Genes for 4,5-PCD, 2,3-PCD into R. jostii RHA1
(22) The ligAB genes encoding S. paucimobilis protocatechuate 4,5-dioxygenase (Noda et al. J. Bacteriol. 1990, 172, 2704-2709) were cloned into a suitable expression vector used for inducible gene expression in Rhodococcus, using a thiostrepton inducer (Nakashima; Tamura Appl. Environ. Microbiol. 2004, 70, 5557-5568), generating a construct containing ligAB.
(23) The praA gene encoding Paenibacillus sp. JJ-1b protocatechuate 2,3-dioxygenase (Kasai et al. J. Bacteriol. 2009, 191, 6758-6768) was cloned into an expression vector to give a construct containing praA.
(24) The two constructs were each transformed into Rhodococcus jostii RHA1 via electroporation.
(25) In order to verify expression of the recombinant genes, cell-free extract was obtained from cultures of R. jostii containing the ligAB gene and R. jostii containing the praA gene after induction with 1 μg/ml thiostrepton. Addition of the extract to solutions containing protocatechuic acid gave rise to a yellow colour in each case, corresponding to the meta-ring cleavage product, and absorbance changes of 0.2-0.45 absorbance units over 60 min at 410 nm and 350 nm for constructs containing ligAB and praA, respectively, corresponding to the literature values for the ring cleavage products for protocatechuate 4,5-dioxygenase (see Noda et al. above) and protocatechuate 2,3-dioxygenase (see Kasai et al. above), respectively.
(26) Assay of Protocatechuate Ddioxygenase Activity
(27) Cultures (5 mL) of R. jostii (ligAB) or R. jostii (praA) were grown for 24 hours at 30° C. in Luria-Bertani broth containing 50 μg/mL chloramphenicol, then induced with 1 μg/mL thiostrepton, and grown for a further 48 hours. Cell free-extract was prepared by centrifugation (microcentrifuge, 13,000 rpm, 5 min), then resuspension of the cell pellets in 75 μL of 20 mM Tris buffer pH 8.0, addition of lysozyme (5 μL or 5 mg/mL), incubation for 1 hour at 37° C., then sonication and centrifugation (microcentrifuge). Protocatechuic acid (2.5 mM, 800 μL) was added, and the solutions monitored at 350 nm for R. jostii (praA), giving absorbance changes of 0.29-0.42 after 60 min and a visible yellow colouration; and at 410 nm for R. jostii (ligAB), giving absorbance changes of 0.23-0.27 after 60 min and a slight yellow colouration.
(28) Metabolite Production—General Methodology
(29) Cultures (10 mL) of R. jostii (ligAB) or R. jostii (praA) were grown for 24 hours at 30° C. in M9 minimal media (6 g/L Na.sub.2HPO.sub.4, 3 g/L KH.sub.2PO.sub.4, 0.5 g/L NaCl, 1.0 g/L NH.sub.4Cl, 2 mM MgSO.sub.4, 0.5 mM CaCl.sub.2) containing 50 μg/mL chloramphenicol and either 0.1% (w/v) vanillic acid or 1.0% (w/v) wheat straw lignocellulose, then induced with 1 μg/mL thiostrepton, and then grown for a further 7-11 days at 30° C., supplementing with 1 μg/mL thiostrepton at 48 hour intervals. Aliquots (1 mL) were removed, centrifuged (13,000 rpm, microcentrifuge, 5 min), and the supernatant extracted into ethyl acetate (1 mL). The ethyl acetate extract was injected onto a C18 Zorbax Eclipse plus (Agilent) reverse phase HPLC column on an Agilent 1200 Series system, and analysed by LC-MS using a Bruker HTC-Ultra ESI mass spectrometer. The HPLC solvents were water/0.1% trifluoroacetic acid (solvent A) and methanol/0.1% trifluoroacetic acid (solvent B). The applied gradient was 5% B for 5 min; 5-15% B over 10 min; 15-25% B for 8 min; and 25-100% B for 19 min, at a flow rate of 1.0 ml min.sup.−1. 2,4-PDCA and 2,5-PDCA were detected by extracted ion analysis for fragment m/z 168.0, in positive ion mode, and were compared with authentic standards for 2,4-PDCA and 2,5-PDCA.
(30) Production of 2,4-PDCA Using Vanillic Acid
(31) Metabolite production was first tested on M9 minimal media containing 0.1% vanillic acid as carbon source and ammonium chloride as the nitrogen source. Extracts from R. jostii (ligAB) grown on M9 media containing 0.1% vanillic acid showed a new peak at retention time 9.8 min with m/z 167.7 (MH.sup.+) and 189.7 (MNa.sup.+) matching the retention time and mass spectrum of authentic 2,4-pyridinedicarboxylic acid. No metabolite production was observed using wild-type R. jostii RHA1 under the same conditions. Maximal 2,4-PDCA production was observed after 5 and 7 days fermentation. 2,4-PDCA production decreased after 10-12 days. The yield of 2,4-PDCA was determined by comparison with a standard curve of authentic material to be 112 mg per litre culture media.
(32) Production of 2,5-PDCA Using Vanillic Acid
(33) Extracts from R. jostii (praA) containing recombinant praA gene grown for 5 days on M9 media containing 0.1% vanillic acid also generated a new metabolite at retention time 10.0 min with m/z 167.7 (MH.sup.+) and 189.7 (MNa.sup.+) matching the retention time and mass spectrum of authentic 2,5-PDCA. Peak 2,5-PDCA production was observed after 5 days. The yield was determined to be 80 mg 2,5-PDCA per litre culture from LC-MS analysis.
(34) Production of 2,4-PDCA and 2,5-PDCA Using Milled Wheat Straw Lignocellulose
(35) The same constructs were then grown on M9 minimal media containing 1% milled wheat straw lignocellulose. Extracts from R. jostii (ligAB) gave the peak at 9.8 min corresponding to authentic 2,4-PDCA. Maximum production of 2,4-PDCA was observed at 5-7 days, and a yield of 90 mg/litre was determined by comparison with a standard curve for authentic 2,4-PDCA.
(36) Extracts from R. jostii (praA) gave the peak at 10.0 min corresponding to authentic 2,5-PDCA. Maximum 2,5-PDCA production was observed at 5 days. A yield of 79 mg/L was determined by comparison with a standard curve for authentic 2,5-PDCA.
(37) Bioreactor Fermentations—General Methodology
(38) Cultures of R. jostii (ligAB) or R. jostii (praA) were grown at 30° C. in 2.5 L M9 minimal media (6 g/L Na.sub.2HPO.sub.4, 3 g/L KH.sub.2PO.sub.4, 0.5 g/L NaCl, 1.0 g/L NH.sub.4Cl, 2 mM MgSO.sub.4, 0.5 mM CaCl.sub.2) containing 50 μg/mL chloramphenicol and either 0.1% (w/v) vanillic acid or 1.0% (w/v) wheat straw lignocellulose or 0.5% (w/v) Kraft lignin (from Billerud Korsnas Ltd, Sweden) in an Electrolab FerMac 3010 bioreactor. Fermentations were induced by addition of 1 μg/mL thiostrepton after 24 hours, and then grown for a further 3-8 days at 30° C., supplementing with 1 μg/mL thiostrepton at 48 hour intervals. After fermentation, cultures were harvested by centrifugation (6,000 g, 10 min), and the supernatant was applied to Amberlite™ IRA900 anion exchange column (100 mL volume), washed with water (200 mL), and then eluted with 0.5 M HCl (800 mL). 15×50 mL fractions were collected, and analysed by UV-vis spectroscopy for the presence of 2,4-PDCA (λ.sub.max 273 nm, ε=3.1×10.sup.3 M.sup.−1 cm.sup.−1) or 2,5-PDCA (λ.sub.max 271 nm, ε=6.3×10.sup.3 M.sup.−1 cm.sup.−1).
(39) Bioreactor Fermentations—Production of 2,4-PDCA and 2,5-PDCA
(40) The R. jostii (ligAB) construct was grown in a 2.5 L bioreactor in M9 minimal media containing 1% wheat straw lignocellulose for 9 days, reaching a maximum OD.sub.600=1.08 after 5 days, and OD.sub.600=0.90 after 9 days. After centrifugation of bacterial cells, the supernatant was applied to an Amberlite™ IRA900 anion exchange column (100 mL volume), washed with water, and then eluted with 0.5 M HCl. The eluted fractions showed absorbance maxima at 273 nm and 230 nm matching those of authentic 2,4-PDCA (ε=3.1×10.sup.3 M.sup.−1 cm.sup.−1), which was eluted in 15×50 mL fractions, with fractions 5 and 6 showing the greatest amount of product. The amount of 2,4-PDA present in the organic extracts after 9 days fermentation was 125 mg/L (estimated by LC-MS analysis). The yield of material after ion-exchange chromatography, based on UV-vis analysis, was 102 mg/L.
(41) A fermentation of R. jostii (ligAB) in M9 minimal media containing 0.5% Kraft lignin over 4 days produced 2,4-PDCA (confirmed by HPLC analysis). Product isolation via Amberlite™ IRA900 anion exchange chromatography resulted in a yield of 53 mg/L 2,4-PDCA.
(42) Growth of the R. jostii (praA) construct in the 2.5 L bioreactor in M9 minimal media containing 1% wheat straw lignocellulose over 9 days, followed by product isolation via Amberlite™ IRA900 anion exchange chromatography, yielded product fractions absorbing at 271 nm matching authentic 2,5-PDCA, with fractions 4 and 5 containing the most product. The amount of 2,5-PDCA present in organic extracts after fermentation for 9 days was 106 mg/L (estimated by LC-MS analysis). The yield of material after ion-exchange chromatography, based on UV-vis analysis, was 65 mg/L.
(43) A summary of the yields of 2,4-PDCA and 2,5-PDCA is provided in Table 1.
(44) TABLE-US-00001 TABLE 1 Yields of 2,4-PDCA and 2,5-PDCA Carbon source for M9 minimal media 0.1% vanillic 1% wheat 0.5% Kraft Construct Product Scale acid straw lignin R. jostii 2,4- 50 mL 112 mg/L.sup.a 90 mg/L.sup.a — (ligAB) PDCA (7 days) (7 days) 2.5 L — 125 mg/L.sup.a 53 mg/L.sup.b bioreactor (9 days) (4 days) 102 mg/L.sup.b R. jostii 2,5- 50 mL 80 mg/L.sup.a 79 mg/L.sup.a — (praA) PDCA (5 days) (5 days) 2.5 L — 106 mg/L.sup.a — bioreactor (9 days) 65 mg/L.sup.b .sup.ayield estimated from LC-MS analysis of the fermentation broth as compared with authentic standard; .sup.byield of product after ion exchange chromatography, calculated using UV-vis absorption. Length of fermentation time indicated in brackets.
Example 2
Synthesis of 2,4-diethyl-2,4-pyridinedicarboxylate
(45) ##STR00020##
(46) 2,4-pyridinedicarboxylic acid (25.38 g; 150 mmol) was added to ethanol (1,300 mL). Aqueous sulfuric acid (2.54 g) was added. The mixture was heated at reflux (about 78° C.) for 15 hours, during which time water was removed from the reaction. The reaction progress was monitored using NMR spectroscopy. After the 2,4-diethyl-2,4-pyridinedicarboxylate had been formed in >97% purity by NMR, the reaction mixture was allowed to cool to ambient temperature and was extracted with 2-methyltetrahydrofuran. The combined organic layers were washed with a saturated aqueous brine solution and deionised water, and dried (MgSO.sub.4). The organics were filtered and the volatiles were removed in vacuo to afford the title compound (24.49 g; 110 mmol; >99% conversion by GC).
Example 3
Synthesis of 2,5-diethyl-2,5-pyridinedicarboxylate
(47) ##STR00021##
(48) 2,5-pyridinedicarboxylic acid (25.08 g; 150 mmol) was added to ethanol (1,800 mL). Aqueous sulfuric acid (1.32 g) was added. The mixture was heated at reflux (about 78° C.) for 58 hours, during which time water was removed from the reaction. The reaction progress was monitored using NMR spectroscopy. After the 2,5-diethyl-2,5-pyridinedicarboxylate had been formed in >97% purity by NMR, the reaction mixture was allowed to cool to ambient temperature and was extracted with 2-methyltetrahydrofuran. The combined organic layers were washed with a saturated aqueous brine solution and deionised water, and dried (MgSO.sub.4). The organics were filtered and the volatiles were removed in vacuo to afford the title compound (26.61 g; 120 mmol; >99% conversion by GC).
(49) General Methodology for the Formation of Copolymers
(50) A 250 mL flange flask with 5 quick-fit ports was used. Stirring was achieved via a magnetic stirrer using a large precious metal stirrer bar. The rates of stirring were gradually decreased from the initial 400 rpm down to 200 rpm to avoid issues as a result of the increasing viscosity of the reaction mixture. All reagents were added to the reactor and warmed to 110 to 130° C. as described below to allow total melting and achieve miscibility. A flow of N.sub.2 gas was applied for 20 minutes to purge the reagents and reactor of oxygen. The temperature was then increased to the desired point as stated below. After a further four hours of very low N.sub.2 flow the gas line was removed, and a vacuum pump turned on, initially at a low vacuum (˜200 mbar) but slowly increased as stated below.
Example 4
Synthesis of 2,4-polybutyrate adipate pyridinedicarboxylate (2,4-PBAP)
(51) ##STR00022##
(52) 2,4-Diethyl-2,4-pyridinedicarboxylate (22.32 g; 100 mmol), diethyl adipate (20.23 g; 100 mmol), 1,4-butane diol (22.53, 250 mmol) and titanium(IV) tert-butoxide (0.77 mL; cat.) were combined. The reaction mixture was heated at 110° C. for 4 hours at atmospheric pressure with stirring at 400 rpm, 180° C. for 17 hours at 200 mbar and 350 rpm, and at 180° C. for 3 hours at 25 mbar and 250 rpm. The copolymer was formed (32.9 g). The .sup.1H NMR spectra for 2,4-PBAP can be found at
(53) The ratio of 2,4-pyridinedicarboxylate:adipate was determined by .sup.1H NMR to be 0.957:1. The molecular weight of the 2,4-PBAP was estimated using end-group analysis, wherein the ratio of end groups to those of the bulk polymer were calculated using .sup.1H NMR to give the number of constitutional repeating units (CRU), which was estimated to be 9.60. One ideal CRU is 421.46 gmol.sup.−1. Therefore, the molecular weight of the 2,4-PBAP was estimated to be 4,044.7 gmol.sup.−1.
Example 5
Synthesis of 2,5-polybutyrate adipate pyridinedicarboxylate (2,5-PBAP)
(54) ##STR00023##
(55) 2,5-Diethyl-2,5-pyridinedicarboxylate (22.32 g; 100 mmol), diethyl adipate (20.23 g; 100 mmol), 1,4-butane diol (22.53, 250 mmol) and titanium(IV) tert-butoxide (0.77 mL; cat.) were combined. The reaction mixture was heated at 110° C. for 4 hours at atmospheric pressure with stirring at 400 rpm, 180° C. for 17 hours at 200 mbar and 350 rpm, and at 180° C. for 3 hours at 25 mbar and 250 rpm. The copolymer was formed (40.62 g). The .sup.1H NMR spectra for 2,5-PBAP can be found at
(56) The molecular weight of the 2,5-PBAP was estimated by .sup.1H NMR using end-group analysis as described for 2,4-PBAP. The ratio of 2,5-pyridinedicarboxylate:adipate was determined to be 0.92:1. The number of CRUs was estimated to be 10.15. One ideal CRU is 421.46 gmol.sup.−1. Therefore, the molecular weight of the 2,5-PBAP was estimated to be 4,278.8 gmol.sup.−1.
Comparative Example 6
Synthesis of Polybutyrate Adipate Terephthalate (PBAT)
(57) Diethyl terephthalate (22.22 g; 100 mmol), diethyl adipate (20.23 g; 100 mmol), 1,4-butane diol (22.73, 230 mmol) and titanium(IV) tert-butoxide (0.77 mL; cat.) were combined. The reaction mixture was heated at 130° C. for 2 hours at atmospheric pressure with stirring at 400 rpm, 180° C. for 2 hours at atmospheric pressure and 400 rpm, 180° C. for 17 hours at 200 mbar and 350 rpm, and at 180° C. for 3 hours at 25 mbar and 250 rpm. The copolymer was formed (40.51 g). The .sup.1H NMR spectra for PBAT can be found at
(58) The molecular weight of the PBAT was estimated by .sup.1H NMR using end-group analysis as described for 2,4-PBAP. The ratio of terephthalate:adipate was determined to be 1.047:1.
(59) The number of CRUs was estimated to be 16.4. One ideal CRU is 420.45 gmol.sup.−1. Therefore, the molecular weight of the PBAT was estimated to be 6,893 gmol.sup.−1.
(60) PBAT is available commercially under a range of trade names. The molecular weight of one particular commercial PBAT was estimated by .sup.1H NMR using end-group analysis as described for 2,4-PBAP. The ratio of terephthalate:adipate was determined to be 0.93:1. The number of CRUs was estimated to be 25.7. One ideal CRU is 420.45 gmol.sup.−1. Therefore, the molecular weight of the commercial PBAT was estimated to be 10,809 gmol.sup.−1. The .sup.1H NMR spectra for commercial PBAT can be found at
Example 7
(61) Thermal Analysis of Polymers Using (STA and DSC)
(62) The thermal stability of cured copolymer was analysed using Simultaneous Thermal Analysis (STA) using a Stanton Redcroft STA 625. Approximately 10-20 mg of copolymer was heated from ambient temperature to 625° C. at a heating rate of 10° C. min-1 under nitrogen. Typically, two distinct decompositions were observed; when present, the first may be the decomposition of end-groups and is thus often small, the second may be the major decomposition of the copolymer backbone. The results can be found in Table 2.
(63) TABLE-US-00002 TABLE 2 STA analysis of polymers Temperature of Temperature of Temperature 5 wt % loss 1.sup.st decomp. of 2.sup.nd decomp. STA Copolymer ° C. ° C. ° C. trace 2,4-BPAP 304.7 — 356.2 FIG. 5 2,5-PBAP 289.1 328 381.5 FIG. 6 Comparative 289.5 — 406.0 FIG. 7 Example 6 Commercial 341.5 — 409.5 FIG. 8 PBAT
(64) The glass transition temperature (T.sub.g) and melting point (T.sub.m) of the copolymers were obtained by Differential Scanning Calorimetry (DSC) analysis using a TA Instruments Q2000 DSC. Indium was used as the standard to calibrate the temperature and heat capacity. Copolymer samples (7-10 mg) were sealed in T zero aluminum hermetic DSC pans. The method was carried out under a constant flow of dry nitrogen of 50 mL/min, at 10° C./min over a temperature range of −80° C. to 250° C. The results can be found in Table 3. The DSC traces can be found at
(65) TABLE-US-00003 TABLE 3 DSC analysis of copolymers T.sub.g1 T.sub.g2 T.sub.m Copolymer ° C. ° C. ° C. 2,4-BPAP −31.7 — — 2,5-PBAP −35.3 41.9 105.9 Comparative −39.5 42.4 134.6 Example 6 Commercial −30.1 45.4 122.2 PBAT
Example 8
(66) Gel Permeation Chromatography (GPC)
(67) The molecular weight (M.sub.n and M.sub.w) and polydispersity (Pd.sub.i) data as generated by GPC can be found in Table 4. GPC was conducted on an Agilent SECurity GPC System 1260 Infinity using THF as the solvent, a polystyrene standard, and a light scattering detector.
(68) TABLE-US-00004 TABLE 4 GPC analysis of copolymers Copolymer M.sub.n M.sub.w Pd.sub.i GPC chromatogram 2,4-PBAP 323.3 1,109 3.4 FIG. 9 2,5-PBAP 2,448 5,186 2.11 FIG. 10 Comparative 5,582 8,615 1.544 FIG. 11 Example 6 Commercial 42,190 113,100 2.680 FIG. 12 PBAT
Example 9
(69) Tensile Strength Measurement
(70) Mechanical properties including tensile strength, elongation at break and Young's modulus of samples are summarised in Table 5. Film samples were prepared by heating about 8 g of copolymer in a fan-assisted oven at 160° C. for 15 min (180° C. for PBAT). The resulting films were cut into standard dumb-bell shapes (60 mm×10 mm). Film thickness was in the region of 1.5-2.0 mm. Tensile studies were conducted in triplicate using an Instron 3367 universal testing machine fitted with 1000 N capacity load cell. The initial grip separation was set at 35 mm and the crosshead speed was 20 mm/min. The results reported were the average of the three measurements (the elongation at break was obtained automatically from the software). Commercial PBAT is a typical elastomer with elongation over 293%. It has the highest tensile strength over 19.5 MPa and good Young's modulus of 100.8 MPa.
(71) TABLE-US-00005 TABLE 5 Tensile strength measurement of copolymers Tensile strength Elongation at break Young's Modulus Copolymer MPa % MPa 2,4-PBAP — — — 2,5-PBAP 2.8 ± 0.4 5.2 ± 0.3 90.6 ± 14.0 Comparative 4.8 ± 0.5 2.3 ± 0.2 269.8 ± 0.2 Example 6 Commercial >19.5 >293.1 100.8 PBAT
(72) The 2,4-PBAP and 2,5-PBAP copolymers produced are soft like that of the commercial PBAT. The expected ratio of PDCA to adipate of about 1:1 has been incorporated into the copolymer. The observed molecular weight of 2,4-PBAP, 2,5-PBAP and comparative example 6 (PBAT) are significantly lower than that of commercial PBAT. This is expected given the relatively small scale on which the copolymerisations were conducted and will be higher in a full scale production process. The NMR data provides an indication of the relative number of constitutional repeating units (CRU) and hence an indication of molecule weight, though the GPC provides more accurate values.
(73) The differences in the data obtained for the copolymers of the invention and the commercial BPAT may be partly attributed to a lack of branching in 2,4-PBAP and 2,5-PBAP.
(74) Those skilled in the art will recognise or be able to ascertain using no more than routine experimentation many equivalents to the specific embodiments described herein. The scope of the present embodiments described herein is not intended to be limited to the above description, but rather is as set forth in the appended claims. Those of ordinary skill in the art will appreciate that various changes and modifications to this description may be made without departing from the spirit or scope of the present invention, as defined in the following claims.
Example 10
(75) Stabilised green waste compost is matured in a composting bin under controlled aeration conditions. Before use, the mature compost is sieved on a screen of 5 mm. The fine fraction forms the inoculum with a total solids content of approximately 50-55% and the volatile content of the total solids is more than 30%.
(76) The standard and control materials are mixed with the inoculum in a ratio of approximately 1 to 1.5 parts of total solids to 6 parts of total solids and introduced into a reactor. These reactors are closed and put into an incubator. The temperature of the reactors is maintained at 58° C. +/−2° C. Pressurised air is pumped through a gas flow controller and blown into the composting vessel at the bottom through a porous plate. During biodegradation, solid carbon of the test sample is converted into CO.sub.2.
(77) The gas leaving each individual reactor is analysed at regular intervals for CO.sub.2 and O.sub.2 concentrations. As the flow rate is continually measured, the cumulative CO.sub.2 production can be determined. The percentage of biodegradation is determined as the percentage of solid carbon of the test compound that is converted into CO.sub.2.