生物降解

u b o f . the conversion of suberin, the biopolymer proposed to be responsible for the waterproon have been reported on the transf provides a signicant opportun value. The suberin content varies bark. The most studied cases are (Betula pendula) which are report up to 40% and 58% wt/wt of the d The widely accepted generic st proposed by Bernards,5 contai complex and heterogeneous, lig aliphatic polymer consisting of neous catalysts to sustainable technologies,8 hydrogenolysis of RSC Advances PAPER Pu bl ish ed o n 06 S ep te m be r 2 01 3. D ow nl oa de d by U ni ve rs ity o f S ci en ce an d Te ch no lo gy o f C hi na o n 24 /0 9/ 20 13 0 1: 08 :0 6. aSchool of Chemistry and Chemical Eng University Belfast, Belfast BT9 5AG, UK. E- qub.ac.uk; g.sheldrake@qub.ac.uk; Fax: +44 bJohnson Matthey Technology Centre, Blount 9NH, UK † Electronic supplementary informati chromatographic analysis of products fr See DOI: 10.1039/c3ra44382e This journal is ª The Royal Society of g nature of bark. Few studies ormation of this material yet it ity due to its low commercial widely in different species of cork (Quercus suber) and birch ed to contain suberin at levels ry extracted bark respectively.4 ructure of suberin, originally ns two primary domains; a nin-like polymer and a largely hydroxycinnamates esteried the bark with heterogeneous precious metal catalysts was employed to depolymerise this biomass source. The use of hydrogenolysis to depolymerise biomass, and specically wood, has been known for many years. Excellent reviews by Sakaki- bara9 and, more recently, Bruijnincx et al.10 report the extensive work using both homogeneous and heterogeneous catalysts to produce a range of monomeric and dimeric aromatic mole- cules. Pepper and co-workers11 have examined, in detail, the heterogeneous metal-catalysed hydrogenolysis of different woods with rhodium on carbon emerging as the preferred catalyst. In all cases, the aromatic products 2-methoxy-4-pro- pylphenol (4-propylguaiacol, 1) and 4-(3-hydroxypropyl)-2- methoxyphenol (dihydroconiferyl alcohol, 2), shown in Fig. 1, New methods in b hydrogenolysis of Mark D. Garrett,a Stephen C and Gary N. Sheldrake*a Hydrogenolysis of bark from three catalysts produces two major prod and the lignin-like regions of su a,u-functionalised species, are pr demonstrate clear advantages o conversion and product selectivity Introduction Petroleum is a non-renewable natural resource and still provides us with the majority of primary chemicals.1 Biomass is the obvious renewable alternative and has the potential to provide both aromatic and aliphatic compounds for a sustain- able chemical industry.2 Although the efficiency and sophisti- cation of bioreneries has improved considerably in the last decade3 the range of commercially viable compounds available from biomass is still limited to a relatively small range of chemical building blocks. The development of efficient tech- nologies for the conversion of biomass to both new and existing platform chemicals continues to be an important challenge. This paper reports a new catalytic hydrogenolysis approach for Cite this: DOI: 10.1039/c3ra44382e Received 4th June 2013 Accepted 5th September 2013 DOI: 10.1039/c3ra44382e www.rsc.org/advances ineering, David Keir Building, Queen's mail: m.garrett@qub.ac.uk; c.hardacre@ 28 9097 4687 's Court, Sonning Common, Reading, RG4 on (ESI) available: 1H-NMR and om the depolymerisation of the bark. Chemistry 2013 iomass depolymerisation: catalytic barks† . Bennett,b Christopher Hardacre,*a Robin Patricka different species of tree using heterogeneous platinum group metal ct streams. Aromatic substituted guaiacols are produced from lignin erin and a range of saturated fatty acids and alcohols, including duced from the polyester regions of suberin. Control experiments catalytic hydrogenolysis over base hydrolysis, both in terms of with glycerol or long chain u-hydroxy fatty acids. Therefore, within bark, the aromatic domains from both suberin and lignin are available to produce aromatic monomers whereas the aliphatic domain of suberin is a potential source of long chain lipids and unusual u-functionalised fatty acids. Previous studies on the depolymerisation of bark to small molecules have concentrated on suberin-rich species of cork and birch6,7 and the characterisation of fatty acids in suberin. This present research has focused on utilising the whole bark by isolating both aromatic and aliphatic product streams from the biopolymer. In general, depolymerisation of suberin has been accomplished by base hydrolysis (e.g.with sodiummethoxide or potassium hydroxide) to cleave the polyester linkages. Within the umbrella of our research on the application of heteroge- View Article Online View Journal were the major products obtained from the lignin of so woods. In this study, the main focus was the application of platinum group metal heterogeneous catalysts in the depolymerisation of the lignin-based sections of bark together with, for the rst time to the best of our knowledge, the release of the fatty acids from the suberin polymer. To understand the inuence of the different proportions of suberin and lignin between wood RSC Adv. Derivatisation for GC-MS analysis RSC Advances Paper Pu bl ish ed o n 06 S ep te m be r 2 01 3. D ow nl oa de d by U ni ve rs ity o f S ci en ce an d Te ch no lo gy o f C hi na o n 24 /0 9/ 20 13 0 1: 08 :0 6. View Article Online species, our study concentrated on sycamore, spruce and cork. This range of barks was chosen to cover both valuable commercial woods, e.g. spruce, which have a signicant waste production from the forestry industry, a fast growing, ubiqui- tous species (sycamore) and a species containing high levels of suberin (cork). Experimental General Hydrogenolysis reactions were carried in a 100 mL Autoclave Engineer mini-reactor. GC-MS was performed using a Perkin- Elmer autosystem XL GC with a Perkin-Elmer Turbomass detector and BP5 column. 1H NMR spectroscopy was performed on a Bruker Avance spectrometer at 300 MHz with TMS as internal standard unless indicated otherwise. Spruce bark was obtained from Borregaard Industries. Sycamore bark was obtained from local trees in Northern Ireland and cork samples were obtained from commercial wine bottles. All bark was pulverised and sieved to <0.2 mm. The material was subse- quently pre-dried at 70 �C for at least 2 weeks in air. The bark samples were then used directly without any solvent pre-ex- traction. Rhodium on carbon (5% Rh, 64% wet) and palladium on carbon (5% Pd, 60% wet) were supplied by Johnson-Matthey. 1,4-Dioxane was supplied by Alfa-Aesar and distilled and doubly deionised water was used in all reactions. All other reagents Fig. 1 Structures of two aromatic products from wood depolymerisation. used for derivatisation and analytical standards were obtained from Sigma Aldrich. Quantitative analysis using 1H NMR spec- troscopy was carried out by dissolving 100 mg of the oil extract from the reaction in deuteriated chloroform (5 mL) and deu- teriated methanol (0.1 mL). To 1 mL of this solution was added 1 mL of an internal standard solution containing 1.0 mg mL�1 vanillin in deuteriated chloroform. Comparison of the integra- tions of the observed sample peaks with the vanillin aldehydic peak enabled quantication. General procedure for hydrogenolysis Initially the autoclave was loaded with catalyst (0.5 g, 64% wet), bark (3.0 g) and 1 : 1 dioxane–water (44 mL). Aer exchange of the head space with hydrogen, the pressure was set to 600 psi H2 pressure and the stirring set to 1000 rpm. Thereaer, the RSC Adv. Results and discussion Hydrogenolysis of dried samples of sycamore, spruce and cork bark at 200 �C and 40 bar hydrogen pressure catalysed by Rh/C and Pd/C resulted in up to 13% weight yield of organic-soluble oils isolated by chloroform extraction from the liquid phase of the reaction. Preliminary analysis of the isolated crude oils by 1H NMR spectroscopy clearly showed the presence of both aromatic and fatty acid products. However, closer analysis revealed multiple peaks due to partially depolymerised lipids and esters. Further hydrolysis of the oil in dioxane–water with sodium hydroxide resulted in a simplied product mixture showing distinctive signals in the 1H NMR spectrum attributed to methylene groups a to free carboxylic acids. The solid residue isolated by initial ltration of the hydrogenolysis reaction was used to determine an approximate degree of depolymerisation Approximately 20 mg of the extracted oil-aer NaOH reux was placed in a solution of 1.25 M HCl in anhydrous methanol (2 mL). This mixture was heated under reux for 4 h under nitrogen aer which the methanol was removed by evaporation using a nitrogen stream. The residue was suspended in water (5 mL) and extracted with chloroform (3 � 5 mL). The organic layer was dried and concentrated using a nitrogen stream to an oily residue which was further dried under vacuum for 2 h. Aer this time the oil was dissolved in anhydrous pyridine (0.3 mL), N,O-bis(trimethylsilyl)-triuoroacetamide (0.5 mL containing 10% chlorotrimethylsilane) was then added. This mixture was heated at 70 �C with agitation for 1 h under nitrogen. Aer cooling, 0.2 mL of a standard of 1 mg mL�1 (trimethylsilyl)- cholesterol was added. This mixture was injected directly into the GC-MS apparatus for characterisation. Quantitative analysis of the fatty acids using GC-MS was accomplished by comparing the retention times with a number of families of compounds namely alkanoic acids, diacids, hydroxyacids and alkanols. The concentration of the compounds present was obtained using the response factors normalised to a (trimethylsilyl)cholesterol standard of the families of standard compounds previously measured. temperature was increased to 200 �C and this temperature was maintained for 4 h. Aer cooling, the reaction was ltered and the lter cake washed with 1 : 1 dioxane : water (3 � 20 mL). The combined ltrates were extracted with chloroform (3 � 200 mL), dried over anhydrous magnesium sulphate and concentrated under vacuum producing an oil. This oil was taken up in 1 : 1 dioxane : water (30 mL) to which was added sodium hydroxide (0.5 g). Aer reuxing for 4 h the mixture was extracted with chloroform (3� 100mL), dried and concentrated to yield a brown oil. While the use of chloroform as extraction solvent is clearly not desirable from a green chemistry perspective, its use in this context is purely for convenience of obtaining clean product solutions for analysis and the devel- opment of alternative separation procedures for larger scale production are under investigation. This journal is ª The Royal Society of Chemistry 2013 of the original bark during the catalytic process. Table 1 summarises the crude yields and level of depolymerisation using three sources of bark and the two heterogeneous catalysts. The dominant aromatic species within the extracted oils were identied as propylguaiacol (1) and dihydroconifery- lalcohol (2) (Fig. 1) using 1H NMR spectroscopy and GC-MS analysis following comparison with pure analytical standards. While minor amounts of other aromatics such as ferulic acid, catechin12 and 3,4-dihydroxybenzoic acid13 were reported in previous bark depolymerisation studies, this is the rst report, to the best of our knowledge, of the isolation of 1 and 2 from bark. Both 1 and 2 have been reported previously as products from the hydrogenolysis of wood and, recently, ethylguaiacol was reported as a product from the hydrogenolysis of lignosulpho- nate.14 The similarity in hydrogenolysis products to wood in this case suggests that the lignin polymer regions within the bark are the origin of these compounds although some contribution from the aromatic regions of the suberin cannot be ruled out. Quantitative analysis to determine the levels of aromatic prod- ucts within the crude isolated oils was performed using 1H NMR spectroscopy. Integration the aldehydic signal of vanillin at 9.73 ppm as an internal standard to compare with the side-chain methylene signals a to the aromatic rings of propylguaiacol and dihydroconiferyl alcohol, at 2.44 ppm and 2.55 ppm respec- tively, enabled calculation of the relative proportions of pro- pylguaiacol (1) and dihydroconiferyl alcohol (2). These results are summarised in Table 2. A signicant catalytic effect is evident by comparing the yields of aromatic products with the control reactions contain- ing no catalyst (Table 2). Furthermore, the low yield of the control reaction of sycamore bark with Rh/C but without hydrogen demonstrated that the effect of the metal catalyst is through a hydrogenolysis reaction and not due, for example, to catalysed hydrolysis. The highest yield for sycamore bark depolymerisation was obtained using 5%Rh/C which gavemore than double the quantity of extractable products compared with 5% Pd/C. With spruce bark, both catalysts showed similar behaviour and at least a 5 fold increase in aromatic products was observed compared with the non-catalysed experiments. However, no correlation could be drawn between the yields of Table 1 Isolated yields from organic extraction after hydrogenolysis of barks and percentage weight reduction of initial bark after hydrogenolysis Catalyst/bark species Oil (wt%) Weight reduction of bark (%) Rh/C Sycamore 9.3 56 Spruce 9.1 65 Cork 11.5 33 Pd/C Sycamore 13.3 50 Spruce 9.2 66 Cork 7.2 38 Table 2 Aromatic yields from hydrogenolysis of sycamore, spruce and cork barks ti o Paper RSC Advances Pu bl ish ed o n 06 S ep te m be r 2 01 3. D ow nl oa de d by U ni ve rs ity o f S ci en ce an d Te ch no lo gy o f C hi na o n 24 /0 9/ 20 13 0 1: 08 :0 6. View Article Online Bark/catalyst Yield (mg) Run 1 Increase w.r.t control (n-fold) Aroma (wt%) Rh/C Sycamore 114 6.3 3.8 Spruce 36 5.3 1.2 Cork 40 3.3 2.0 Pd/C Sycamore 63 3.5 2.1 Spruce 45 6.6 1.5 Cork 26 2.2 1.3 Control reactions Sycamorea 18 — 0.6 Sycamoreb 24 — 0.8 Sprucea 6.8 — 0.2 Corka 12 — 0.6 a Hydrogenolysis carried out under same reaction conditions except with catalyst. This journal is ª The Royal Society of Chemistry 2013 aromatic products and the suberin contents within each of the barks (cork (40%) > sycamore (26%) > spruce (<10%)) reported previously.15 Therefore, as the lignin contents are similar for all three substrates (at about 25%) it seems likely that the aromatic products originate mostly from the lignin polymer of the barks rather than the aromatic domain of the suberin. The yields of aromatic products from the bark substrates reported here greatly exceed the aromatic product yields reported in the majority of simple hydrolysis depolymerisation studies. For example, ferulic acid is the most commonly reported aromatic product from suberin hydrolysis, usually at levels of <1%,16 and with and without Rh/C or Pd/C Run 2 cs Yield (mg) Increase w.r.t control (n-fold) Aromatics (wt%) 81 4.5 2.7 45 6.6 1.5 46 3.8 2.1 46 2.5 1.5 63 9.3 2.1 38 3.2 1.9 ut catalyst. b Hydrogenolysis carried out without hydrogen but with Rh/C RSC Adv. occasionally as high as 4.3%.17,18 Although lower, the yields of aromatic products from these tree barks are much closer to the levels of 9–10% of total aromatic products from the hydro- genolysis of whole wood biomass.11 Previous studies on the hydrolytic depolymerisation of suberin from various bark species reported a variety of long chain (C12–C28) fatty acids and esters and long chain alcohols (Fig. 2).4,15 The characteristic peaks of such lipids were also evident in the 1H NMR spectra of the crude oil extracts from our hydrogenolysis experiments. Alkanoic acids in the oil were characterised by the presence of a triplet at about 0.9 ppm corresponding to the terminal methyl groups. u-Hydroxy- alkanoic acids and alkanols were seen to be present from the CH2OH triplet at 3.6 ppm. Quantication of the total lipid and fatty acid content (excluding the alkanols) was determined using a vanillin internal standard and comparing the integra- tion of aldehydic singlet in the 1H NMR spectrum to the distinctive peak at 2.25 ppm from the CH2 a to the fatty acid/ ester carbonyl group (Table 3). The results from Table 3 demonstrate that hydrogenolysis with Rh/C or Pd/C results in improved yields of characterisable prod- ucts from all three bark species when compared with the unca- talysed control reactions. The suberin domain of spruce bark is most easily depolymerised with an increase, with respect to the control reaction, of greater than 90% with either catalyst and, signicantly, a 158% increase over the control using Pd/C in one particular run. This result is extremely encouraging as spruce bark in particular is a high volumewastematerial from industrial wood processing. Comparing the two catalysts, Rh/C has the greatest overall catalytic activity with yields increasing by more than twice those found for Pd/C for both sycamore and cork. For Pd/C reactions, the yield increase with respect to the uncatalysed control reaction improves from cork to sycamore to spruce. This may suggest that the level of depolymerisation of bark to lipids is not linked to the percentage suberin in each bark as reported suberin levels in these barks increase from spruce (<10%) to Fig. 2 Typical lipid families reported from hydrolysis reactions.15 Table 3 Lipid yields from hydrogenolysis of sycamore, spruce and cork barks with Run 1 s o RSC Advances Paper Pu bl ish ed o n 06 S ep te m be r 2 01 3. D ow nl oa de d by U ni ve rs ity o f S ci en ce an d Te ch no lo gy o f C hi na o n 24 /0 9/ 20 13 0 1: 08 :0 6. View Article Online Bark/catalyst Yield (mg) Increase w.r.t control (%) Lipid Rh/C Sycamore 87 81 2.9 Spruce 84 133 2.8 Cork 52 63 2.6 Pd/C Sycamore 60 25 2.0 Spruce 93 158 3.1 Cork 38 19 1.9 Control reactions Sycamorea 48 — 1.6 Sprucea 36 — 1.2 Corka 32 — 1.6 a Hydrogenolysis carried out under same reaction condition except with RSC Adv. sycamore (26%) to cork (40%). However, such results demonstrate the widespread accessibility of different barks of changing biopolymer composition to depolymerisation using catalysed hydrogenolysis. Studies on the hydrolytic depolymerisation of suberin-rich barks such as cork and birch produced high yields of fatty acids per weight of bark. Yields of 12.6% and 16.0% for cork,19 and up to 26.0% for birch bark7 have been reported. Although the levels of fatty acids per weight of bark achieved in this study are signicantly lower than conventional base hydrolysis the hydrogenolysis results, herein, demonstrate that the Rh and Pd catalysts can fragment the aliphatic region of the suberin structure as well as the aromatic regions. These yields may be compared with the baseline hydrolysis using our reac- tion conditions in a mixture of dioxane–water and sodium hydroxide. This reaction resulted in lower yields of fatty acids and without Rh/C or Pd/C Run 2 (wt%) Yield (mg) Increase w.r.t control (%) Lipid (wt%) 96 100 3.2 69 92 2.3 46 44 2.3 69 44 2.3 78 117 2.6 35 9.3 1.8 ut catalyst. This journal is ª The Royal Society of Chemistry 2013 (2.0 %wt lipid/wt bark) and insignicant amounts of aromatics. This proof of concept of catalytic hydrogenolysis of barks clearly demonstrated the effect of the heterogeneous catalyst for the catalytic depolymerisation of suberin. More detailed speciation of the lipids produced using cata- lysed hydrogenolysis was obtained from the hydrogenolysis of sycamore bark, using Rh/C as a typical example. The extracts were derivatised to produce corresponding methyl esters from the carboxylic acids and silyl ethers from the alcohol groups to aid volatility and then analysed using GC-MS. To compare