Brønsted Acid Ionic Liquid Catalyzed Depolymerization of Poly-(3-hydroxybutyrate) to 3 ‑ Hydroxybutyric Acid: Highly Selective and Sustainable Transformation in Methyl Isobutyl Ketone and Water-Containing Phase-Separable Reaction Media

: Poly-(3-hydroxybutyrate), PHB, is a bacterial poly-ester in industrial demand as a biodegradable alternative to fossil-derived nondegradable plastics. Moreover, apart from being used directly as a bioplastic, valorization of PHB to its monomer building blocks and other value-added chemicals is feasible but less explored. In this study, Brønsted acid ionic liquid (BAIL) catalyzed depolymerization of PHB was investigated as a highly selective route to 3-hydroxybutyric acid, 3-HBA. The hydrolysis of PHB to 3-HBA was performed in a biphasic solvent medium composed of methyl isobutyl ketone (MIBK) and water, where the organic phase had dual roles as an efficient medium for dissolution of the polymer and as solvent for the monomeric products, which were enriched in this phase after cooling, with the Brønsted acid ionic liquid (BAIL) catalyst partitioned into the aqueous phase for facile recycling. The effects of reaction parameters, including the temperature, types of IL in terms of cations and anions, and the amount of water and IL, were studied to assess the yield of 3-HBA. Furthermore, protic acids such as sulfuric acid, methanesulfonic acid, and p -toluenesulfonic acid ( p -TsOH) were also applied for comparison as acid catalysts for the hydrolysis of PHB to 3-HBA. Among the tested catalysts, the ILs containing the p -TsO − as anion as well as p -TsOH alone were found to be highly selective in promoting hydrolysis to 3-HBA, with complete depolymerization of PHB at >90% yield of 3-HBA in 4 h at 120 ° C using a BAIL with sulfobutylated 1-methylimidazolium as the cation component and p -TsO − as the anion ([ImSO 3 H + ][ p -TsO − ]). Although the use of p -TsOH as the sole catalyst also yielded efficient PHB hydrolysis with high reaction rates, it had a disturbing effect on the biphasic MIBK − water system by forming a single-phase reaction mixture at high 3-HBA yields, obstructing the recoveries of the products as well as the catalyst. In contrast, the biphasic reaction mixture remained intact when using IL as catalyst, which allowed facile and efficient separation of the product from the catalyst. Both the 3-HBA and the [ImSO 3 H + ][ p -TsO − ] IL were recovered in high purity, the latter after applying a solvent extraction scheme based on ethyl acetate, whereby the recoveries of 3-HBA and IL reached ≈ 90%. The compositions of the synthesized ILs and the progress of the hydrolysis process, as well as the purity of the recovered product, were confirmed by NMR analysis. This sustainable approach to selective hydrolytic transformation of PHB into 3-HBA using a recoverable acidic IL catalyst in a biphasic solvent media of aqueous methyl isobutyl ketone hence resulted in efficient product separation and catalyst recovery.


■ INTRODUCTION
Microbial polyesters, such as polyhydroxyalkanoates (PHAs), are produced by many microorganisms as a means of storing energy.−3 Bacterial PHAs and their derivatives are considered as biocompatible plastics, 4 that are biodegradable due to their ability to decompose under both aerobic and anerobic conditions. 5,6−9 Hence, PHAs have a vast potential to replace petroleum derived nondegradable plastics and to reduce plastic waste and prevent the formation of nondegradable marine microplastic.Due to their interesting properties, PHAs are explored in numerous applications including drug delivery, agriculture, textile, paper coating, energy materials, implants, automobiles, nonwoven fabrics, biosensors, food coating and packing, molded goods, adhesives, etc. 7−10 A diversity of metabolic pathways are involved in the synthesis and accumulation of PHAs in microbial cells, resulting in the formation of polymers with a variety of monomeric compositions, including homo-and copolymers. 11,12Critical parameters affecting the overall economy of PHA production are the availability of suitable feedstocks, alongside the cost of fermentation, recovery, and other downstream processes. 12A variety of feedstocks has been used to produce PHAs by bacterial fermentation.7 Notwithstanding this multitude of feedstocks, the annual worldwide production of these environmentally friendly polymers has reached only about 50000 tons, which is only a tiny fraction of the production of fossil plastics, which in 2019 exceeded 450000000 tons.18 Poly(3-hydroxybutyrate) (PHB) appears to be the biopolyester most commonly targeted for transformation of carbon feedstocks by bacteria, microalgae, cyanobacteria, and even transgenic plants. 2 Utilization of the polymers harvested from cells is almost invariably oriented toward processing and use as such, suffering from variations in molecular weight, copolymer composition, and crystallinity inherent in bioprocessing of feedstocks with varying quality, 19 factors that have substantial influence on the physical and chemical properties of the final products.Yet, attention has only recently been drawn to PHB and other PHAs as starting materials in biorefinery schemes after depolymerization, 20 and in the synthesis of composites and low molecular weight organics of industrial demand.21,22 By noncatalytic and solventless thermal depolymerization PHAs can be transformed to unsaturated carboxylic acids such as crotonic acid (CA), isopropyl-2-CA, 3-butenoic acid, and 2-pentenoic acid, in addition to other organics such as γ-valerolactone, as well as low molecular weight dimers and trimers.23−25 In this thermally induced pyrolytic process (≈300 °C), repetitive βelimination facilitates scission of the polymeric chains and allows for the formation of lower molecular weight polymeric chains with unsaturated end groups, which subsequently are converted to unsaturated carboxylic acids.23,24 In the case of PHB, the pyrolysis method not only enables the depolymerization with >90% selectivity for CA, but it also facilitates a continuous product separation through distillation.26 Alongside solventless thermal processing, high temperature hydrolysis and acid−base catalytic approaches have also been applied to convert PHAs to small organics, with PHB being more extensively studied compared to other PHA polymers.22,27,28 Such studies have shown that PHB can be depolymerized to C 4 -platform molecules such as 3-hydroxybutyric acid (3-HBA) and/or CA by a hydrothermal approach, with and without the addition of acid or base catalysts (Scheme 1).25,27 Both 3-HBA and CA have been valorized by hydrothermal treatment to industrially valuable polymeric materials (CA-vinyl acetate copolymer), pharmaceuticals, and plastic precursors, as well as gaseous hydrocarbons such as propylene.22,26,29 In the absence of additives or catalysts, hydrolytic depolymerization of PHB typically requires temperatures above 200 °C.These rather harsh conditions are accompanied by incomplete conversion of the polymer and low selectivity toward 3-HBA and CA, 30 whereas catalysts such as sulfuric acid, sodium hydroxide, sodium acetate, magnesium oxide, and hydroxides allow the depolymerization to be performed under milder conditions, with complete conversion of PHB and high selectivity for 3-HBA and CA.27,30 Besides inorganic acids or bases, ionic liquids (ILs) have also been used as catalytic solvents for the depolymerization of PHB to 3-HBA and CA, where both acidic and basic ILs facilitate the formation of desired products with high selectivity.Ionic liquids (ILs) are considered as special nonaqueous and ionic solvent media relevant in many applications, including their use as a solvent and/or catalyst in various organic conversion, as well as for dissolution of biopolymers for valorization to precious C 2 −C 5 platform molecules and aromatics.31−33 For base catalysis, our group has reported on alkaline ionic liquids such as 1-ethyl-3-methyl imidazolium acetate as combined catalyst and solvent media for highly selective depolymerization of PHB to CA achieving >90% selectivity and yield for CA.34 Brønsted acid ILs (BAILs) have also previously been utilized by Song and co-workers for the solvolysis of PHB into 3-HBA and methyl 3-hydroxybutyrate (Me-3-HB) in aqueous and methanolic media, respectively. 35−37 The conversion of PHB in aqueous medium reached 98% with 1-methyl-3-(3-sulfopropyl)-imidazolium hydrogen sulfate, [HSO 3 -pmim][HSO 4 ] as acidic IL catalyst. Hoever, the authors did not mention concomitant formation of CA, which could be suspected since the reactions were performed at temperatures up to 170 °C.Neither did they °C with recycling of the ILs without significant loss of the activity.Solvolysis of PHB with BAIL is hence demonstrated to be relatively milder and selective in methanol compared to an aqueous medium.
Once formed in the acid-catalyzed depolymerization of PHA, 3-HBA remains unstable in the acidic aqueous reaction mixture and undergoes subsequent dehydration to CA, especially at elevated temperatures, reducing the selectivity of the process 30 (Scheme 2).Additionally, because of the drawback of operating a pressurized process at high temperature, corrosion of equipment, difficulties in additive recycling, and waste generation, the formation of 3-HBA through acidic hydrolysis in a neat aqueous medium is considered suboptimal in terms of both economic efficiency and environmental sustainability.In this report, we propose for the first-time application of BAILs for the hydrolysis of PHB to 3-HBA with high selectivity with methyl isobutyl ketone (MIBK) as cosolvent to enhance the product selectivity.MIBK is among the preferred solvents in biorefining and is applied as a lowcost extraction medium in biomass fractionation as well as in the synthesis of low molecular weight organic moieties by catalytic depolymerization of biorenewables, such as sugars, chitosan, etc. 38,39 Li et al. also efficiently applied a biphasic MIBK−H 2 O reaction medium for the selective extraction of organosolv lignin from agriculture waste such as corncob, corn stalk, rice husk, and rice straw, achieving up to 91.9% extraction of lignin. 40−43 In this case, selective liquid−liquid extraction of these furfurals into the MIBK phase allowed for their synthesis with high selectivity.In practice, this in situ extraction of 5-HMF or FF from the aqueous to organic phase after their formation diminishes further interaction with the acid catalyst in the aqueous medium, hence reducing the risk of undesirable side reactions such as self-polycondensation.In the case of the acid-catalyzed transformation of chitin or chitosan, MIBK was also used as an efficient extractant for the separation of levulinic acid from the aqueous phase, which further facilitates the reuse of acidic ILs as a catalyst in consecutive catalytic cycles. 44,45n this report, we exploit the efficient swelling and dissolution of PHB at modestly elevated temperatures in a MIBK−water solvent mix for a selective acid-catalyzed hydrolysis of PHB into 3-HBA, catalyzed by BAIL.The reaction was studied under various combinations of reaction parameters such as composition of ILs in terms of types of cations and anions, amounts of water and IL, catalyst concentration, and reaction temperature.The progress of the PHB hydrolysis reaction was monitored by 1 H NMR spectroscopy.The fortunate formation of a biphasic reaction mixture, where the product and the IL catalyst are found in separate phases, facilitated the recovery of both the product and solvent and allowed the BAIL to be recycled for subsequent catalytic cycles.
Synthesis and Characterization of Brønsted Acid Ionic Liquids (BAILs).The BAILs were synthesized in two reaction steps following a procedure previously reported in the literature. 33As shown in Scheme 3, the first reaction step involved zwitterion formation by sulfobutylation using equimolar amounts of 1,4butanesultone with various amines or triphenylphosphine.The zwitterions were subsequently converted to BAILs by reactions with equimolar amounts of various protic acids.The compositions and purities of the synthesized BAILs were confirmed by 1 H NMR analysis with spectra acquired at 298 K using a Bruker (Billerica, MA, U.S.A.) DRX-400 spectrometer with a 9.4 T magnet, corresponding to Larmor precession frequencies for protons of 400.2 MHz.The data obtained were further processed with TopSpin 4.0.7 software from Bruker.
The relative acid strengths of the BAILs and protic acids applied in the synthesis and hydrolysis of PHB was determined as the Hammett acidity function (H 0 ) 47 with 4-nitroaniline as indicator using a VWR UV-3100PC Spectrophotometer (VWR, Radnor, PA, U.S.A.) according to a slightly modified previously reported procedure. 33rior to UV−vis spectroscopic measurements, 0.001 mol of BAILs or protic acids were mixed with 5 mL of a 0.1 mM solution of 4nitroaniline in absolute ethanol followed by stirring of the mixture for 1 h at room temperature.The absorbances (A max ) of the alcoholic solution of BAIL and protic acids, as well as a reference solution of 4-Scheme 2. Acid-Catalyzed Dehydration of 3-Hydroxybutyric Acid to Crotonic Acid Scheme 3. Synthesis of BAILs a a BY 3 = 1-methylimidazole, triethylamine, pyridine, or triphenylphosphine; HX = sulfuric acid, methanesulfonic acid, or p-toluenesulfonic acid.
nitroaniline, were measured.The measurements were carried out in a quartz cuvette with a 10 mm path length, where the wavelength of 370 nm (λ max ) was taken as the maximum absorbance.The absorbance values were used to evaluate the Brønsted acidity of the BAIL and protic acids in terms of the Hammett acidity function (H 0 ), where pK(I) is the pK a value of protonated 4-nitroaniline in an aqueous solution, while [I] and [IH + ] are the molar concentrations of the unprotonated and protonated forms of 4-nitroaniline, respectively.Acid-Catalyzed Hydrolysis of Poly(3-hydroxybutyrate) to 3-Hydroxybutyric Acid.Hydrolysis of PHB catalyzed by Brønsted acid ionic liquid or protic acid was carried out in 5 mL pressure resistant screwcap glass vials, which after capping were fixed to a custom-built agitation unit designed to tumble the vials end-overend at ≈ 30 rpm.This agitator was located in the oven of a modified HP 5890 gas chromatograph (Hewlett-Packard, Palo Alto, CA, U.S.A.).If not otherwise noted, the base reaction compositions, found after a series of scouting experiments, consisted of 0.2 g of PHB, 0.5 mM of protic acid or BAIL, 0.21 g of water (11.7 mmol; five times molar excess based on the monomer units in PHB) and 1.738 g (17.4 mmol) of MIBK, which were mixed prior to the experiment.The vials were then attached to the tumbling device after the oven had been preheated to isothermal temperatures of 80, 100, 120 or 140 °C, respectively.After the reaction had completed, each vial was removed from the oven and allowed to cool at room temperature to allow the phases to separate.
The 3-HBA and CA formed in the reactions was determined by 1 H NMR as previously described, 34,46 against a calibration curve in CDCl 3 or DMSO-d 6 with methanol as internal standard.Prior to the NMR analyses, stock solutions of 50 μL of methanol in 10 mL of CDCl 3 or DMSO-d 6 were prepared to establish calibration curves for quantification of 3-HBA in the actual reaction mixtures.The required amounts of 3-HBA were mixed with 0.5 mL of these stock solutions, and the mixtures were analyzed by 1 H NMR. The ratios of the integrated signals for the CH 3 groups of 3-HBA and methanol were used to draw calibration curves (Figure S1; figures with prefix S are found in the Supporting Information).The amount of CA was calculated by means of the methyl group using the 3-HBA calibration curve.At predetermined time intervals, ≈35 mg aliquots of the reaction mixture were collected, mixed with 0.5 mL of stock solution, and analyzed by NMR to determine the yields of 3-HBA and CA.For protic acid-catalyzed processes, the reaction mixtures were shaken thoroughly, and weighed quantities of the reaction mixtures were used for NMR analyses with DMSO-d 6 as a solvent.Analysis of the organic phase of samples from three separate runs with the base conditions listed above at 120 °C for 4 h resulted in measured 3-HBA concentrations of 91.96, 87.97, 90.02 (90.0 ± 2.00; RSD = 2.2%) and CA concentrations of 7.13, 6.56, 7.11 (6.93 ± 0.32; RSD = 4.7%).
Attempts to Model the Reaction Kinetics.Experiments aimed at determining the kinetics parameters of the overall reaction in the biphasic reaction media were undertaken with p-TsOH•H 2 O and [ImSO 3 H][p-TsO] IL as catalysts at 100, 120, 140, and 160 °C over a time span of 30 to 240 min, with the experiment at 100 °C extended up to 720 min.Samples were withdrawn at regular intervals, and the amounts of 3-HBA and CA were determined by NMR as described above.The modeling and optimization software MODEST was used to elaborate the kinetic models and for processing the data, as previously described. 34,48,49ecovery of Brønsted Acid Ionic Liquid and 3-HBA.The biphasic reaction mixture used to hydrolyze PHB at 120 °C for 4 h with [ImSO 3 H][p-TsO] IL as catalyst was used in the experiments to recover the BAIL and 3-HBA.Prior to carrying out the recovery experiments, complete conversion of PHB was verified by NMR analysis.After completion of the reaction, the vials were kept at room temperature for 1 h to ensure complete phase separation.The upper phase, containing MIBK and 3-HBA, was carefully suctioned off and transferred to a round-bottom flask, followed by washing the remaining water phase with fresh portions of MIBK (three times 2 mL), which were combined with the MIBK phase first drawn off.The MIBK was then removed by gentle rotary evaporation at 40 °C to recover the 3-HBA from the collected fraction.Subsequently the remaining traces of organic solvents and CA were removed by high vacuum.The sublimation characteristics of CA thereby facilitated its efficient separation from 3-HBA under mild conditions.Next, the aqueous bottom phase containing BAIL was also treated by rotary evaporation, followed by applying high vacuum at 40 °C to remove traces of water from the [ImSO 3 H][p-TsO] IL.Three aliquots of 5 mL of ethyl acetate were used to wash the recovered BAIL, followed by drying, first by rotary evaporation and thereafter in high vacuum.The recoveries of 3-HBA and BAIL were calculated using eqs 2 and 3, respectively, while the purities were confirmed by 1 H NMR analysis.
%recovery of 3 HBA recovered 3 HBA(g) 100 theoretical amt of 3 HBA(g) = × (2) ■ RESULTS AND DISCUSSION Synthesis and Characterization of Brønsted Acid Ionic Liquids.The synthesis of BAILs was carried out by an equimolar reaction of 1,4-butanesultone with various amines (1-methylimidazole, triethylamine, or pyridine) or triphenylphosphine in the presence of protic acids, yielding the BAILs in Figure 1.The purity and composition of the synthesized ILs were confirmed by 1 H NMR spectroscopy (Figure S2).The acid strengths in terms of the Hammett acidity function were also measured spectrophotometrically, with the results shown in Table 1.
As shown in Table 1, the absorbance (A max ) value for the unprotonated 4-nitrotroaniline was 0.241, which was decreased as the indicator was protonated by the BAILs and the protic acids.The observed H 0 values for the protic acids were 1.87 or 1.88, whereas the H 0 values for the BAILs ranged between 1.90 and 2.03, indicating that the protic acids are slightly more acidic than the BAILs. 37cid-Catalyzed Hydrolysis of PHB to 3-HBA with BAILs and Protic Acids.Considering the Hammett acidity parameter (H 0 ) values in Table 1, the protic acids showed acid strengths only slightly higher than those of the BAILs.This acid strength similarity prompted us to investigate the catalytic abilities of the protic acids MSA, p-TsOH, and H 2 SO 4 for the hydrolysis of PHB prior to examining the ILs as catalysts.The protic acid tests consisted of heating reaction vials containing mixtures of PHB, water, MIBK, and protic acid to 120 °C for varying times from 1 to 4 h.As shown in Figure S4a, the PHB remained essentially unreacted when H 2 SO 4 was tested as catalyst.In a separate control experiment, it was observed that PHB was soluble in MIBK at 120 °C and formed a viscous gel when the solution was cooled to room temperature.A dense aqueous phase had formed at the bottom of the vial since PHB is essentially insoluble in water and the mutual solubility of water and MIBK is limited (0.103 ± 0.0027 wt.% water in MIBK and 0.338 ± 0.0096 wt.% MIBK in water at 25 °C).50 With MSA as catalyst, PHB was steadily depolymerized as the reaction time increased from 1 to 4 h, and no solids were seen in the reaction mixtures after 3 and 4 h of reaction (Figure S4b).The conversion of PHB occurred even more efficiently with p-TsOH compared to MSA, as the solid PHB completely disappeared after 1 h, forming a reaction mixture that remained transparent as the reaction time continued to 4 h (Figure S4c).The yields of the expected products from the hydrolysis of PHB were measured quantitively by 1 H NMR using calibration curves obtained with methanol as the internal standard and DMSO-d 6 as the solvent.Corresponding NMR spectra and yields of possible products are summarized in Figure 2 and Figure S3.The attempt to use H 2 SO 4 as hydrolysis catalyst for PHB resulted in formation of merely 2.8% of 3-HBA after 4 h, with most of the polymer remaining unreacted.On the other hand, when MSA and p-TsOH were used as catalysts for the hydrolysis of PHB under otherwise identical reaction condition, the yields of 3-HBA reached 67% and 94% after 4 h, respectively (Figure 2).Alongside the formation of 3-HBA, its dehydration product CA was also formed during the acid-catalyzed hydrolysis of PHB.A negligible amount of CA was found in the H 2 SO 4 -catalyzed process liquor, presumably because production of the precursor 3-HBA failed.The yields of CA reached 3.4 and 5.2% when MSA and p-TsOH were used as the catalysts, respectively (Figure 2).Although H 2 SO 4 , MSA, and p-TsOH had practically identical Hammett acidity parameters (H 0 ), their catalytic activities in this PHB hydrolysis scheme were very different.Both MSA and p-TsOH have also previously been used as versatile catalysts in valorization of biopolymers (e.g., lignocellulose, polysaccharides etc.) 51−53 and recycling of poly(ethylene terephthalate).54 Studying the Mechanism of the Acid-Catalyzed Depolymerization of PHB in Aqueous MIBK Media.The mechanism for the acid-catalyzed hydrolysis of PHB was elucidated by using a reaction mixture processed at 120 °C for 1 h with MSA as acid catalyst, analyzing the reaction mixture obtained after cooling by heteronuclear multiple bond correlation (HMBC) and two-dimensional (2D) correlation NMR.
The 1 H− 13 C HMBC 2D NMR spectrum in Figure 3a shows numerous correlation peaks between the carbonyl carbons (−C�O) and aliphatic protons such as in −CH and −CH 2 groups in the MSA-catalyzed reaction mixture.Correlation signals between carbon atom in the −C�O group and protons in −CH and −CH 2 groups for PHB were obtained at chemical shifts 5.10/168.96and 2.53/168.96ppm (red rectangles; color coding in Figure 4), respectively, while correlation peaks for the same groups in 3-HBA were obtained with chemical shifts of 3.97/172.84and 2.26/172.84ppm (blue rectangles, Figure 3a), respectively.The chemical shifts for the carbonyl carbon atoms of PHB and 3-HBA were confirmed by acquiring 13 C NMR spectra of these separately (Figure 3b) and comparing those with the spectra obtained from the reaction mixture.In addition, as shown in the 2D NMR analysis, smaller proton peaks for unknown chemical entities at 2.28, 3.96, and 5.096 ppm were also observed where all these protons showed correlation peaks (black rectangles) with the same carbonyl carbon of chemical shift 170.23 ppm.Moreover, the protons with chemical shifts of 2.51 and 5.096 ppm also showed additional correlation peaks with another carbonyl carbon with a chemical shift of 171.49ppm (pink rectangles).Among the peaks obtained for the unknown entities, the protons with chemical shifts 2.28 and 3.96 ppm have chemical shifts similar to the protons of the −CH and − CH 2 groups in 3-HBA, respectively (cf. Figure S3).Likewise, the protons at 2.51 and 5.096 ppm also have chemical shifts similar to those of the proton of the −CH and −CH 2 groups in     gradually smaller size with 3-hydroxybutyrate groups at their termini, as well as nonterminated oligomers (Figure 4b,c).Subsequently, these depolymerized species undergo terminal hydrolysis, leading to the formation of 3-HBA.This continuous depolymerization facilitates the sequential conversion of PHB to 3-HBA (Figure 4a).It is worth mentioning that the correlation peaks mentioned above at 5.10/168.96and 2.53/ 168.96 ppm belong to the oligomeric entity of PHB without 3hydroxybutyrate group, shown in Figure 4c, which are also formed during depolymerization.A similar mechanism was observed in our previous publication on base catalyzed thermal depolymerization of PHB to CA, where the polymer was initially depolymerized to crotonyl group terminated and nonterminated oligomers, which were eventually depolymerized to CA. 34 The higher catalytic activity of p-TsOH compared to MSA (Figure 2), called for an extension of the temperature range in the PHB hydrolysis experiments using p-TsOH as catalyst to investigate the product selectivity between 3-HBA and CA, since the formation of CA by dehydration of 3-HBA is temperature-dependent. 30Additional experiments were therefore conducted by varying the reaction time at 100, 140, and 160 °C, with the corresponding NMR spectra shown in Figures S7 and S8.At 100 °C, a substantial amount of PHB remained undissolved after 4 h.Most of these solids had dissolved after 6 h, but it took 12 h to obtain a completely transparent reaction mixture (Figure S5a).In contrast, at 120 °C, transparent reaction mixtures were obtained when the reaction preceded from 30 min to 4 h (Figure S5b).When processing of PHB was performed at 140 and 160 °C, the reaction mixtures became transparent within 30 min to 4 h.
As shown in Figure 5 with corresponding NMR spectra in Figures S7 and S8, the amounts of 3-HBA and CA varied significantly in the reaction media as the reactions progressed at temperatures from 100 to 160 °C.At 100 °C, hydrolysis of PHB was slow, with 81.7% of the monomer transformed to 3-HBA in 6 h and negligible formation of CA.As the reaction continued for six more hours, the yield of 3-HBA increased to 93.6%, but in the reaction mixture now contained 5.7% CA.Increasing the reaction temperature from 100 to 120 °C resulted in significantly increased reaction rates, where PHB was practically entirely hydrolyzed in 2 h to 96.3% 3-HBA and 3.5% CA, summing to 99.8% of the charged monomer units.However, as the reaction was allowed to continue for 2 h, the amount of yield of 3-HBA decreased to 94.1% with a corresponding increase of CA due to the dehydration of 3-HBA (Scheme 2).At 140 °C the reaction rate increased further with 93.2% yield of 3-HBA after 30 min, but this was reduced to 75.9% by further processing up to 4 h with CA now obtained at 23.2% yield.The spontaneous formation followed by dehydration of 3-HBA was observed at 160 °C where the 86.4% yield of 3-HBA obtained after 30 min was reduced to 36.2% after 4 h, accompanied by a 61.7% yield of CA due to dehydration.Hence, as the reaction temperatures increased, there was a corresponding increase in the rate of PHB hydrolysis but also in the competing reaction to CA. Best yield of 3-HBA was obtained with two h of reaction at 120 °C.The NMR spectra showing increase in the amount of CA with temperature and time are shown in Figure S8.
Solubility and Formation of Multiphase Reaction Mixtures.The formation of a biphasic system was expected since the reaction mixture contained MIBK and an excess of water with limited mutual miscibility (see above).Without additional components capable of homogenizing the reaction mixtures, biphasic systems will also exist in the entire investigated reaction temperature range, since the solubility of MIBK in water remains essentially constant at a mole fraction of 0.025 to 0.03 whereas the mole fraction of water in MIBK varies from ≈0.2 to 0.3 as the temperature is increased from 100 to 160 °C. 50owever, at 120 °C, the mixture became homogeneous and transparent after one hour of reaction, resulting in 85.6% yield for 3-HBA.The mixture also remained homogeneous when the reaction proceeded further to two and four hours with 3-HBA yields > 90%.On the contrary, at 140 and 160 °C, biphasic reaction mixtures were obtained for all reaction times, except for 30 min at 140 °C.To investigate the phase behavior of MIBK/water containing 3-HBA or CA, these expected reaction products were added individually at room temperature at concentrations chosen to match the final compositions of the reaction mixtures obtained after complete conversion of 0.2 g of PHB to 3-HBA or CA, or their combination.As expected, MIBK and water alone led to the formation of a biphasic mixture (Figure S6a).The two-phase system also remained after p-TsOH was added, despite p-TsOH being miscible with both MIBK and water (Figure S6b).When 3-HBA was added to the reaction mixture, followed by shaking, it turned into a single-phase system (Figure S6c).Addition of more water led to phase separation, with the MIBK-rich phase forming the upper layer above a mainly aqueous phase (Figure S6d).An experiment was thereafter carried out with the reverse order of adding p-TsOH and 3-HBA.To our surprise the MIBK and water phases remained separated after addition of 3-HBA, but the system turned into a single phase after addition of p-TsOH.This demonstrates that, despite the limited miscibility of water and MIBK, the interaction of 3-HBA with p-TsOH facilitated the formation of a homogeneous reaction mixture, whereby water became more available for the hydrolysis of the PHA, which was dissolved in the MIBK phase.To gain insight into the interactions between p-TsOH and 3-HBA which led to the formation of single-phase reaction mixtures, the required amounts of 3-HBA (between 0.011 and 0.077 g) were added to five separate NMR tubes where 0.02 g of p-TsOH•H 2 O had been dissolved in 0.45 mL of DMSO-d 6 , in order to establish a series of solutions with 3-HBA:p-TsOH• H 2 O mole ratios 1:1, 2:1, 3:1, 5:1, and 7:1, which were subjected to 1 H NMR analysis.The peak for the acid proton of dry p-TsOH (not shown) has a chemical shift above 11 ppm.However as shown in Figure 6, its monohydrate gave rise to an intense and shielded acid proton peak at 7.18 ppm, due to the equilibrium between H + and H 2 O with H 3 O + .As the quantity of 3-HBA in the composition was increased, the acidic proton peaks of p-TsOH•H 2 O were gradually deshielded and broadened, which was accompanied by a reduction in peak intensity.An equimolar ratio of 3-HBA and p-TsOH•H 2 O gave a chemical shift of 7.26 ppm for the acidic proton peak of p-TsOH, a value that gradually increased with increasing amounts of 3-HBA, reaching 7.43 ppm at a molar ratio of 7:1 (Figure 6).
Similarly, the 1 H NMR spectra of the reaction mixtures obtained at 100 and 120 °C in MIBK/water showed identical shifts for the peak of the acidic proton in hydrated p-TsOH as the amount of 3-HBA was gradually increased in the reaction mixtures.The reaction mixture obtained after processing for 2 h at 100 °C gave rise to a peak at 3.69 ppm for the acidic proton, which was shifted downfield as the reaction further proceeded to 4, 6, and 12 h since the amount of 3-HBA increased in the reaction mixture (Figure S8).Moreover, at 120 °C, the chemical shift for the acidic proton increased until 2 h, where 3-HBA yield reached 96.3%.However, after 3 and 4 h, the chemical shift values ceased to increase since the yield of 3-HBA had not started to decrease significantly (Figure S8).This 1 H NMR analysis reveals that the presence of increasing amounts of 3-HBA in the reaction mixture led to strong interactions between 3-HBA and the hydrated p-TsOH species.This enhances the miscibility of water in the MIBK phase, facilitating the formation of a homogeneous reaction mixture boosting the PHB hydrolysis rate.
As described above, the formation of homogeneous phases was restricted to the reaction mixtures obtained at 100 and 120 °C, whereas the reaction mixtures formed two-phase systems when the reaction temperatures were increased to 140 and 160 °C.As shown in Figure S8, at a reaction temperature of 140 °C, the acidic proton of the hydrated p-TsOH in the reaction mixture underwent a downfield shift as the reaction time increased from 30 min to 1 h.This trend toward higher chemical shift values suggests a stronger deshielding effect on the acidic proton, indicative of increased interactions with other chemical species.Notably, this extended reaction time led to a 3-HBA yield of >90%.However, as the reaction proceeded further, the amount of CA increased steadily in the reaction mixture, accompanied by an increased amount of water due to dehydration of 3-HBA (Scheme 2).Meanwhile, the broadened signal for the acidic proton did not increase and remained practically identical as the reaction time was increased from 1 to 4 h.This absence of a change in the signal of the acidic proton suggests that excess water formed during the reaction did not remain in the MIBK phase to participate in the interactive scheme of p-TsOH, 3-HBA, and water.Instead, it must have been driven out of the MIBK phase to form a biphasic system with the bottom phase consisting mainly of water due to its limited miscibility with MIBK and the other reaction components.Formation of a biphasic reaction mixture was not caused merely by the presence of CA formed by dehydration of 3-HBA, as a control experiment (not shown) formed a homogeneous phase when p-TsOH and CA (at an amount equivalent to the 3-HBA monomer in the charged PHB) were added to MIBK and water at their initial reaction mixture mass ratio.This hints that the biphasic reaction mixtures formed at 140 °C and above are caused by the release of additional water due to dehydration of 3-HBA, which is also supported by the observation above where the homogeneous phase formed by MIBK, 3-HBA, and p-TsOH turned biphasic after addition of an excess amount of water (Figure S6d).At 160 °C, the highest of the tested temperatures, CA and water formed even faster, which resulted in the formation of two-phase systems at all reaction times (Figure S8).Additionally, it was observed that resonances corresponding to the acidic proton of p-TsOH in all reaction mixtures obtained at 160 °C had consistent chemical shifts.This can be attributed to the separation of excess water formed during the reaction, as previously explained, which does not interfere with the interaction between p-TsOH, CA (and/or 3-HBA), and water.
Hence, p-TsOH efficiently catalyzed the hydrolysis of PHB to 3-HBA in high yield at 100 and 120 °C whereas subsequent dehydration of 3-HBA to CA limited the yield of 3-HBA at higher reaction temperatures.However, although 3-HBA was formed in high yield under stable reaction conditions at 100 and 120 °C, the formation of a homogeneous mixture after the reaction did not facilitate the separation of the reaction components from the catalyst and the solvent, which is important to achieve good reaction economy.
Depolymerization of PHB Using Brønsted Acid Ionic Liquids (BAILs) as Catalysts.Acidic hydrolysis of PHB to 3-HBA was next carried out with BAILs as acid catalysts with MIBK and water as solvent media, as for the protic acids, to solubilize the polymer and to act as a hydrolyzing agent, respectively.Initially, the hydrolysis of PHB was attempted with [ImSO 3 H][p-TsO] IL as catalyst at 120 °C in the 1 to 4 h time range.The reaction progress was monitored each hour for the formation of 3-HBA and CA, the dehydrated byproduct of 3-HBA.As in the processes catalyzed by p-TsOH • H 2 O, the [ImSO 3 H][p-TsO] IL also acted as an efficient catalyst for the hydrolysis of PHB, with a yield of 3-HBA reaching 92% after 4 h.The 3-HBA formed was also found to be sufficiently stable at the applied temperature, as merely 7.1% of CA was produced (Figure 7).However, compared to protic acid catalysis with p-TsOH (Figure 5), using the IL caused a more extended lag in 3-HBA formation.With p-TsOH, 85.6% 3-HBA yield was achieved after 1 h at 120 °C, whereas by IL catalysis, it reached only 25%.We attribute this to a beneficial miscibility of p-TsOH•H 2 O in both water and MIBK even at room temperature, whereas the [ImSO 3 H][p-TsO] IL is highly miscible in water but has negligible solubility in MIBK.In addition, as described above, when the amount of 3-HBA in the reaction mixture increases gradually, it promotes mixing of p-TsOH and water into the MIBK phase, and as a consequence, the rate of hydrolysis increases.The solubility of the [ImSO 3 H][p-TsO] IL in MIBK can be assumed to increase at elevated temperatures, which should further facilitate the hydrolysis of PHB as the reaction proceeds, similarly to p-TsOH catalyzed hydrolysis discussed above.
Yet, in contrast to the homogeneous phases obtained with p-TsOH as the catalyst, a biphasic reaction mixture persisted with [ImSO 3 H][p-TsO] IL, as shown in Figure 8, even after 3-HBA had been formed in high yield.In this case, the MIBK phase comprising 3-HBA and CA remained in the upper phase while the lower phase contained the IL in water.It is therefore apparent that relatively high concentrations of ILs combined with their salting-out properties not only enhances the phase separation, but also shifts the extraction coefficient of 3-HBA toward the organic phase.Hence, the IL catalyzed process with MIBK as solvent not only accomplished efficient dissolution of PHB with subsequent conversion to 3-HBA in the optimal reaction temperature range, but it also facilitated the separation of the product from the IL catalyst.
Extending the Study to a Wider Temperature Range.The hydrolysis of PHB catalyzed by [ImSO 3 H][p-TsO] IL was further examined at 100, 140, and 160 °C, with yields of 3-HBA and CA at these temperatures shown in Figure 7 and the corresponding 1 H NMR spectra of the reaction mixtures in Figures S9 and S10.At 100 °C, a significant lag was observed with only 2% yield of 3-HBA after 90 min of reaction.The rate of 3-HBA production thereafter increased to 37.8% after 6 h.However, the yield of 3-HBA after 12 h of reaction was only 61.6%, which is significantly lower than the reaction catalyzed by p-TsOH, which reached 93.6% in the same time.As described above, the higher miscibility of p-TsOH in both water and MIBK compared to its IL analogue [ImSO 3 H][p-TsO] facilitated a more efficient depolymerization under otherwise identical reaction conditions.Moreover, solubility of the PHB polymer at 100 °C was limited and the reaction thus progressed in a nonhomogeneous mixture.At the higher reaction temperatures of 120, 140, and 160 °C, the depolymerization of PHB was more efficient with yields of 3-HBA and CA similar to those obtained with p-TsOH.
Attempts to Determine the Reaction Kinetics.A study intended to chart the reaction kinetics based on the yields of 3-HBA and CA was attempted by first fitting the experimental data to a simple model representing hydrolysis directly from the PHB polymer to 3-HBA coupled to a dehydrating side reaction to CA.However, this model failed because it could not take into account the lags in the production of 3-HBA, which are evident at the lowest reaction temperatures in Figures 5 and 7. A more elaborate model was thereafter devised, intended to encompass the swelling and dissolution of the solid polymer that must take place prior to the actual depolymerization, accompanied by a gradual shift in catalyst focus from in-chain cleavage to terminal monomer detachment.Although this model was capable of following the lagged curves more faithfully, it could not produce reliable reaction rate constants or activation energies for the first two steps of the reactions.In real settings, the initial steps will be strongly dependent on the particle size, molecular weight, copolymer compositions, and crystallinity of the PHB polymer.A kinetics model would therefore be of limited utility, and we therefore refrain from presenting these modeling data.Influence of Water and Ionic Liquid Catalyst Concentrations.After the yields of 3-HBA and CA had been studied as a function of varying reaction temperature, the parameters next in turn to be investigated were the amounts of water and IL.Their influence on the conversion of PHB and the selectivity toward 3-HBA and CA were examined by using two or eight equiv of water with respect to the monomer content in the PHB, in addition to the five equiv of water that had been used in the experiments up to this point.The results are shown in Figure 9a.As noted above, complete conversion of PHB with yields of 92.0% 3-HBA and 7.1% CA was obtained in 4 h at 120 °C when 5 equiv of water was used during the reaction.When the amount of water was decreased to two equiv, both the completeness of the conversion of PHB and the yields of the depolymerized products showed the same pattern of increasing as the reaction progressed from 1 to 4 h.However, the yields of 3-HBA and CA after 4 h were only 66.2% and 4.9%, respectively.Since water is a reactant in the hydrolysis reaction, a decrease of water in the reaction mixture by a factor of 2.5 prevented an efficient hydrolysis of PHB.When the amount of water was instead increased from five to eight equiv, the reaction was also less efficient, with the yield of 3-HBA reaching only 67.7 % after 4 h of reaction, most likely caused by reduced acidity of the BAIL through dilution, leading to a decrease of its catalytic activity.Moreover, additional water decreases the solubility of PHB in the reaction mixture, contributing to the lag that slows the overall reaction.
To study the influence of the amount of IL catalyst, experiments with varying amounts of [ImSO 3 H][p-TsO] were conducted.As shown in Figure 9b, when 10 mg of IL was used, merely 2.8% of 3-HBA was obtained in 4 h at 120 °C with nearly all of PHB remaining unreacted.However, as the amount of IL was increased to 50, 100, and 200 mg, the yields of 3-HBA also increased accordingly to 66.9, 86.0, and 92.0%, respectively, under identical reaction conditions.This was accompanied by a concomitant increase in the amount of CA.This dehydration of 3-HBA is likely not due to catalytic action of [ImSO 3 H][p-TsO] IL, but merely caused by higher concentrations of 3-HBA being available to become dehydrated during a larger fraction of the reaction time.
Probing Different Brønsted Acid Ionic Liquids.As described above, the catalytic power of the protic acids for hydrolysis of PHB varied, despite similar acid strengths (H 0 ).In view of this, a natural experiment extension was to synthesize ILs with varying anion and cation compositions, cf. Figure 1, to be evaluated as catalysts in the hydrolysis of PHB.The yields shown in Figure 10a reveal that all ILs comprising p-TsO − as anion combined with different cations had high and similar efficacies for catalyzing the hydrolysis of PHB with high yields of 3-HBA, i. (IL6; 2.4%).An acid-catalyzed hydrolysis of PHB in a mixed solvent of MIBK and water requires that the polymer, the water acting as a reagent, and the actual acid catalyst, i.e., the proton or the cationic part of the BAIL, are present in the same phase.The highly different catalytic activities of the three [ImSO 3 H]-based BAILs with different anions shown in Figure 10a (p-TsO − in IL4, CH 3 SO 3 − in IL5, and HSO 4 − in IL6) mirror well the differences in activities of the corresponding Bronsted acids, as seen in Figure 2. Since all the investigated acids have practically the same acidity (Table 1), these observations are best explained by different abilities of the Bronsted acids and their corresponding [ImSO 3 H]-based BAILs to partition their catalytic entities, i.e., their protons or the acidic sulfobutylated 1-methylimidazolium cations, into the MIBK-rich phase containing the PHB.Due to the requirement of local electroneutrality, cations cannot transfer from their aqueous environment into the MIBK phase, where the polymer is partitioned, without being accompanied by a balanced amount of anions.Efficient transfer of acidic catalytic entities into the MIBK phase therefore depends on the solubilities of the anions in either phase or their ability to form ion pairs with better solubility in MIBK.
As discussed above, there is a limited solubility of water in MIBK, and in a biphasic system, any entity that binds water strongly will shift the equilibrium of water away from MIBK.Although the sulfonic acid moiety of each of the three anions will have a strong association with water, it is obvious that their overall hydrophilicities and amphiphilic properties differ greatly, with the p-toluenesulfonate ion being the least hydrophilic and most amphiphilic, as opposed to the hydrogen sulfate ion, with the methanesulfonate ion in an intermediate position.The differences in catalytic power in this system can therefore be explained by the hydrophilicity of the anions, which is related to their Setschenow constants describing their efficiency of promoting salting-in or salting-out of nonelectrolytes in binary solvent systems where water is one of the components. 55Hyde and co-workers extended the Setschenow concept by linearizing and normalizing the constants to the salt concentrations, which results in a single number, D norm , that is used to explain the relative power of anions in effectuating salting-out (positive values) or salting-in (negative values) in aqueous two-phase systems. 56 2).This order is inversely related to their distribution from aqueous solutions into the predominantly organic MIBK phase.The hydrogen sulfate ion will moreover contribute to the salting out of the PHB polymer and its neutral reaction products to the MIBK phase and retain water in the aqueous phase, preventing it from distributing into the MIBK phase.This makes the hydrolysis of a relatively hydrophobic polymer such as PHB in a two-phase system quite different from the reported successful depolymerization of lignocellulose by sulfuric acid or BAILs with hydrogen sulfate anions, 57 where the hydrophilic wood fibers are associated with a layer of water on the surface, attracting the acid catalyst to the surface and thereby enhancing its activity.
Reagent Recycling.As mentioned above, the use of [ImSO 3 H][p-TsO] IL as a catalyst resulted in a two-phase system after the depolymerization reaction had completed, which made it possible to separate the upper MIBK phase with the product from the lower aqueous phase containing the [ImSO 3 H][p-TsO] IL catalyst.Recoveries of the IL and 3-HBA after complete hydrolysis of PHB were assessed based on the [ImSO 3 H][p-TsO] IL catalyzed experiments at 120 °C for 4 h.Following the distillation approaches under vacuum, the 3-HBA and IL were recovered at 89 and 94% yield from the reaction mixture, both obtained at high purity (Figure S3).The catalytic power of the recovered IL in the PHB hydrolysis process was explored by recycling the recovered five times, mixing the recovered IL with a new batch of PHB, water, and MIBK and performing the reaction at 120 °C for 4 h.As shown in Figure 10b, the yields of 3-HBA and CA were stable at around 90% and 6%, respectively, with 83% of the originally charged IL recovered at high purity after five times reuse in the hydrolysis of 3-HBA.
Green Process Evaluation.The Process Mass Intensity (PMI) estimated by two different procedures as well as the three "green chemistry metrics" energy economy (ε coefficient), environmental factor (E), and the combined effect of both (ξ) were determined for the depolymerization reaction of PHB using [Bmim] p-TsOH IL as catalyst at 120 °C yielding 94% of the main monomer product and high levels of recycled solvent and catalyst, indicating that the described process is energy-efficient and environmentally friendly when compared to hydrolysis reactions of poly(ethylene terephthalate) (PET), which is the closest analog reaction we could find.A detailed account of these calculations is found in the Supporting Information.

■ CONCLUSIONS
Hydrolytic depolymerization of PHB to 3-HBA was carried out with high recovery using Brønsted acidic IL as catalysts in a mixed MIBK−water solvent media, where the MIBK facilitated the efficient dissolution of PHB and the subsequent product extraction.Alongside the ILs, the corresponding protic acids H 2 SO 4 , p-TsOH, and MSA were also evaluated as catalysts.In spite of the nearly identical acid strengths of the protic acids and Brønsted ILs with different cations and anions, the p-TsOH and ILs with p-TsO − anions gave the highest activity compared to other ILs and protic acids with yields of 3-HBA ranging from 80 to 95% when processed at 120 °C for 4 h.Yields with [ImSO 3 H][MSA] IL and MSA were 38.5% and 67% of 3-HBA, respectively, whereas <10% yield of 3-HBA was obtained with H 2 SO 4 and the corresponding ILs with HSO 4 − anions.Both p-TsOH and [ImSO 3 H][p-TsO] IL gave low reaction rates at 100 °C with 94% and 62% yields of 3-HBA after 12 h, respectively.This is explained by the temperaturedependent solubility of PHB in MIBK.By increasing the reaction temperature to 120, 140, and 160 °C, the rates of hydrolysis of PHB and dehydration of 3-HBA to CA both increased, which led to decreased 3-HBA yields and selectivity at the highest temperatures tested.In the IL-catalyzed process, 5 equiv of water with respect to PHB monomer units gave the highest yield of 3-HBA, whereas 2× and 8× excesses led to decreased yields.The yield of 3-HBA processed at 120 °C for 4 h increased from 2.8% to 92% as the amount of BAIL catalyst was increased from 10 to 200 mg.NMR analysis disclosed that the depolymerization of PHB to 3-HBA proceeded through the formation of 3-HBA-terminated oligomeric PHB intermediates.Despite similar catalytic activities of p-TsOH and [ImSO 3 H][p-TsO] IL reaching >90% yield of 3-HBA under otherwise identical reaction conditions, a single-phase reaction mixture was obtained with p-TsOH, whereas the reaction mixture formed in the BAIL-catalyzed process was biphasic.The biphasic reaction mixture allowed 3-HBA to be selectively extracted from the MIBK phase, and the [ImSO 3 H][p-TsO] IL to be recovered from the water phase.This provided for efficient recovery of both the product and catalyst at high purities, with 89 and 94% recoveries of 3-HBA and IL, respectively.The recovered IL was recycled for five consecutive catalytic cycles, where ∼90% yield of 3-HBA was consistently obtained.Highly selective valorization of PHB to 3-HBA is hence achieved under industrially feasible reaction conditions by a highly sustainable process using a recoverable catalyst in low-cost solvent media facilitating product separation.

Figure 2 .
Figure 2. Yields of 3-hydroxybutyric acid (filled bars) and crotonic acid (stacked open bars) for depolymerization of PHB catalyzed the protic acids p-TsOH, MSA, and H 2 SO 4 at 120 °C for reaction times varying from 1 to 4 h.

Figure 3 .
Figure 3. (a) 1 H− 13 C HMBC 2D NMR of the reaction mixture of hydrolysis of PHB to 3-HBA catalyzed by MSA at 120 °C for 2 h and (b)13 C NMR spectra for spectra for 3-HBA, PHB polymer, and the mixture analyzed in (a), as indicated.

Figure 5 .
Figure 5.Yields of 3-hydroxybutyric acid (3-HBA) and crotonic acid (CA) for the hydrolysis of PHB catalyzed with p-TsOH at different temperatures and reaction times.Reaction conditions: 0.2 g of PHB, 0.5 mM p-TsOH, 0.21 g of water, and 1.738 g of MIBK.

Figure 9 .
Figure 9.Yields of 3-hydroxybutyric acid (3-HBA) and crotonic acid (CA) for [ImSO 3 H][p-TsO] IL catalyzed hydrolysis of PHB at 120 °C, showing the influence of the amounts of (a) water and (b) of [ImSO 3 H][p-TsO].All reactions were composed of 0.2 g of PHB in 1.738 mL of MIBK, with water and [ImSO 3 H][p-TsO] IL as follows: In (a) the [ImSO 3 H][p-TsO] IL was kept constant at 200 mg, with water loadings of 0.084, 0.21, and 0.336 g.In (b) the water was kept constant at 0.21 g, with[ImSO 3 H][p-TsO] IL added at 10, 50, 100, and 200 mg, and the reaction was carried out for 4 h.

Table 1 .
Hammett Acidity Parameters (H 0 ) Measured for BAILs and Protic Acids a a [I] is given in percent of 4-nitroaniline with balance of [IH + ] to 100%.b [PySO 3 H][p-TsO] was not soluble in ethanol.