Combination modified adhesives using strongalkali-degraded soybean proteins利用强碱降解大豆蛋白制备复合改性胶黏剂Hao GU, Zhenhua GAO1, Haiyan TAN, GU JiyouMaterial Science and Engineering College, Northeast Forestry University, Harbin 150040, China顾嗥, 高振华, 谭海彦, 顾继友东北林业大学材料科学与工程学院，哈尔滨 150040Abstract:With decreasing of petroleum resource and increasing environmental concerns, soy-based adhesives will become the leading product of wood industry due to their abundant and renewable predominance. But it has not been widely used because of some disadvantages, mainly the poor water resistance. GPC, DSC and plywood evaluation were employed to characterize the degraded soybean proteins (DSP, treated in the presence of 9wt% sodium hydroxide at 90o C for 2-3.5 hours), the wood adhesives by blending DSP with glyoxal, urea-formaldehyde (UF) resin, melamine-formaldehyde (MF) resin, phenol-formaldehyde (PF) resin and phenol- formaldehyde oligomer. Results indicated that degradation in the presence of strong alkali not only destroyed completely the advanced structures of soy proteins but also resulted in the chain cleavages of the larger-molecular polypeptide, by which obtained a low-viscosity DSP solution suitable for preparing wood adhesive with molecular weight ranged from 282 to 3404. DSC analysis confirmed that the DSP can be crosslinked by glyoxal, UF resin or MF resin, and the curing temperatures of adhesives by blending DSP with PF resin and PF oligomer were from 138 to 182o C. All the wood adhesives prepared using DSP can be used for non-water-resistant applications, but only the DSP-MF blend can be used as water-resistant adhesive.Keywords: soy protein; adhesive; strong alkali degradation; blending; copolymerization;摘要：伴随着石油资源的逐步枯竭以及人们对环境保护的认识日益增强，大豆蛋白胶黏剂以其丰富、可再生的原材料资源以及低廉价格无疑会成为今后木材胶黏剂市场的主导。
但耐水性差的缺陷却限制了这种胶黏剂应用范围，也因此使其至今未能得到广泛的应用。
实验采用GPC、DSC等手段结合胶合板压制，对在90℃和9wt%氢氧化钠存在下降解大豆蛋白及其与乙二醛、UF树脂和MF树脂、PF树脂、PF树脂的低聚物共混，或者与苯酚（P）和甲醛（F）共聚制得的多种改性胶黏剂进行表征，结果表明：降解使大豆蛋白的大分子肽链断裂，高级结构破坏，得到在分子量在282-3404之间、适于制备木材胶黏剂的低黏度产物；随着降解时间延长，产物中大分子量组分含量和黏度逐渐降低，但甲醛反应性能力明显增加；DSC测试表明降解大豆蛋白能够与乙二醛、UF树脂、MF树脂等发生交联固化反应，降解大豆蛋白与PF树脂共混或与PF低聚物共聚制得的胶黏剂固化温度在138℃至142℃之间，低于与UF或者MF树脂复合共混得到的胶黏剂；由降解大豆蛋白制备的胶黏剂均满足室内普通胶要求，而只有含MF树脂的复合胶可达到耐水胶要求。
关键词：大豆蛋白；胶黏剂；强碱降解；共混；共聚Because the biomass resource is resourceful and renewable, more and more attentions were paid to the R&D and highly utilization of biomass resource in recent years. Soybean is primarily an industrial crop widely cultivated in the world for oil and protein, which can not only be used as food resource but also non-food applications such as adhesives for wood and paper, binders in coatings and paints, textile fibers, foams for fire extinguishers, plastics and lubricants for1Corresponding to Zhenhua Gao, gao_zhenhua@substituting for petrochemicals. According to FAO estimates over 240 million tones of soybean will be produced worldwide in 2008. With each ton of crude soybean oil, approximately 4.5 tons of soybean meal (protein content 44%) is produced [1]. Soybean protein fractions were mainly composed of around 52% 11S (a sedimentation constant where S stands for Svedberg units), 35% 7S, 8% 2S and 5% 15S; except the 2S fraction that consists of relatively low molecular mass in the range of 8000–20,000, other fractions in the order of 150,000-600,000 [2], resulting in poor water solubility or lower solid content of the soybean dispersion/solution. In addition of compact globular structure with an internal atomic packing density of 75% [3], soybean protein is undesired for application in wood adhesives.In recent years the modifications of soy-protein based adhesives for wood composites focused on improving their adhesion strength, water resistance, industrial applicability, and reducing their price. Common approach used by most researchers is to modify soy protein with denaturation agents such as alkali [4-5], enzyme [4, 6], urea [7-8], guanidine hydrochloride [7,9], and detergent (sodium dodecyl sulfate and sodium dodecylbenzene sulfonate) [10]; these modifications could improve the bond strength and water resistance to some extent but not to the desired because they just unfold a small quantity of secondary and tertiary structure of soybean proteins [11].Therefore, a new technological concept "degrading soy protein in strong alkaline then blending with polymer or copolymerizing modification for wood adhesives" is proposed in current study. In this case the secondary and tertiary structure of soybean protein was completely destroyed, the macromolecular peptide chain would be also degraded appropriately in the presence of strong alkaline. The treatment can decrease the viscosity, improve the solubility and increase the number of reactive groups of the degraded soybean proteins, and those can be used to make wood adhesive with desired performances for various requirements.1 Experiment1.1MaterialsSoy protein isolate (SPI), with 92.6wt% of protein, purchased from Harbin Hi-Tech Soybean Food Co., Ltd. Other chemicals employed in this study, such as phenol, phosphoric acid, sodium hydroxide and formaldehyde, were purchased from local chemical companies and used without further treatments. Urea-formaldehyde (UF) resin was prepared in our lab. with F/U molar ratio of 1.16 and solid content of 53.6%. Melamine-formaldehyde (MF) resin was prepared with F/M molar ratio of 1.49 and solid content of 54.5%. Phenol-formaldehyde (PF) resin was prepared with F/P mole ratio 2.1 and solid content 42.4%. PF oligomer was prepared by reacting the mixture of phenol, formaldehyde and NaOH at 25-30o C for 48h.Birch veneers (420mm × 420mm, 1.6mm thick) for preparing plywood were purchased from local plywood factory in Harbin.1.2 Experimentations1.2.1 Degraded soybean proteins in strong alkalineHeated 100g of 9wt% NaOH solution to 90-92o C, gradually added 60g SPI powder, and then maintained at 90-95o C for 2-3.5 hours; finally cooled the liquid complex to room temperature for characterization or adhesive preparation. The product was brown transparent without sediment. 1.2.2 Formulations of DSP adhesive by blendingThe detained formulations of each DSP adhesive were listed in Table. When prepared adhesives from J-A to J-F, the pH value of DSP was adjusted to 5.4-5.7 using 85wt% phosphoric acid with agitation. As for adhesives J-G and J-H, the DSP solution was used after degradationwithout pH adjustment.Tab.1 Some formations of DSP adhesives by blendingID J-A J-B J-C J-D J-E J-F J-G J-H AdhesiveDSF content /g 80 80 80 80 80 80 80 80Glyoxal content /g 0 12 0 12 0 12 0 0UF content /g 0 0 24 24 0 0 0 0MF content /g 0 0 0 0 24 24 0 0PF resin content/g 0 0 0 0 0 0 17.5 0PF oligomer content/g 0 0 0 0 0 0 0 22.21.2.3 Molecular weight and distribution of DSP by GPCAgilent 1100 Series GPC-SEC Analysis System (Agilent Company, USA) was employed. The HPLC Columns is 79911 GP-104 PL gel (molecular weight range: 4K-400K) combined with 79911 GP-103 PL gel (molecular weight range: 1-40K); the DSP samples were 0.5%wt% aqueous solution; mobile phase was water with the flow rate of 1mL/min; differential detector. Ten mono-dispersed polyethylene glycols with certain molecular weight were used to calibrate molecular weights.1.2.4 Formaldehyde reactivity of DSPFormaldehyde reactivity, or the formaldehyde reaction ability, was expressed as the weight of formaldehyde in milligrams which was consumed by every gram of soy protein in the DSP solution, tested as follows: add about 40g (W1) of DSP and about 25g (W2) of 37.2% formaldehyde into three-neck bottles with stirring and reflux, adjusted pH value of mixture between 8.6 and 8.7 with phosphoric acid (W3), and then reacted at 85o C for three hours. Finally, measured the residual formaldehyde content (F%) in the reacted mixture according to Chinese Standard GBT 14074-2006 (hydroxylamine hydrochloride method), and calculated Formaldehyde reactivity = 1000 × [37.2% × W2 - (W1 + W2 + W3) × F wt%] / (0.375 × W1).1.2.5 Curing characteristics of DSP adhesives by DSCThe curing characteristics of DSP adhesives were measured in DSC 204 (NETZSCH Company, Germany) under following conditions: heating rate 2.5, 5, 7.5, 10 and 15o C/min from room temperature to 250o C. The Activation energies were calculated according to Kissinger equation [12].1.2.6 Bonding properties of DSP adhesives by plywood evaluationBonding properties of DSP adhesive were evaluated by the bond strength and durability of 3-layer birch plywood panels. Plywood panels were prepared under following conditions: glue consumption 360g/m2 (double bondlines, liquid basis), pre-pressing at 0.8MPa for 5min, hot pressed at 140o C and 1.2MPa for 4.5min. Dry strength and 4h boiled wet strength were tested according to Chinese standards GB/T 17657-1999.2 Results and discussionBecause the soy proteins have compact globular structure with the majority of polar groups and non-polar groups were wrapped inside the globular particles [3], only some polar groups on the surface of soy-protein globular particles could attach to the polar substrate (adherent) and form bonding joint via adsorption. The joints boned by adsorption mechanism were easily damaged under wet condition, resulted in poor water resistance of traditional soy protein based adhesive.Soybean proteins degraded under strongly alkaline condition (pH value of solution> 14) and high temperature (>90o C) could not only completely destroyed intermolecular linkages (mainly hydrogen bonds, disulfide bonds, salt bond and ionic bond between the peptide chain, which built up the advance structures of globular proteins) and fully stretched the separated peptide chains, but also made parts of the primary structures (peptide bonds) hydrolyzed and decrease the molecular weight of peptide chain and solution viscosity. Strong alkali-degraded soybean proteins were different from the alkali-denatured soybean protein that was treated under gentle conditions (pH value usually around 10 and the temperature below 60o C). Alkali-denatured soy proteins still had lots of advanced structure remaining and only a few primary structures might be damaged, resulting in very poor solubility and the solid content of adhesive (protein basis) usually lower than 15wt% [13]. As a water-soluble wood adhesive, the solid content should be not less than 30wt%, otherwise the curing or bonding required more drying time and, therefore, increased the costs and reduced productivity of wood composites.In order to increase the solid content and bond strength and decrease viscosity of soy protein based adhesive, soybean proteins were degraded at 90-92o C in the presence of 9wt% sodium hydroxide for various times. The DSP solutions obtained were brown transparent without sediment, with very low viscosity (32.5-80.5 MPa-s) but higher solid protein content (37.5wt%). Soy proteins were generally composed of about 52% 7S globulin and 35% 11S globulin with molecular weight ranged from 150000 to 60000 [2]. GPC analysis indicated that the DSP solutions had 5 GPC peaks (M1, M2, ..., M5), as shown in Figure 1, corresponding to molecular weight respectively at 3404, 2377, 1803, 863 and 282 that were all far below the molecular weight before degradation. The molecular weight decreased sharply implied that soy proteins were fully degraded. The key component of DSP solution correlated to the M1 GPC peak with molecular weight of 3404 and GPC area percentage between 43.9-50.5% (as shown in Table 2). The GPC curves of each degraded soybean protein for various times were very similar but the GPC peak areas varied to some extent with degrading time increased, as indicated in Table 2. For instance, as the degradation time increasing, the peak area of main GPC peak M1 decreased gradually from 50.5% to 43.9%.Fig.1 GPC spectra of degraded soy proteins with various timesFig.2 DSC curves of adhesive J-A and J-B with various heating rates During the degradation of soybean protein in the presence of strong alkali, one macromolecular polypeptide chain decomposed into two smaller polypeptide chains via the hydrolysis of one amide group into one terminal amino and one terminal carboxyl group. As shown in Table 2, formaldehyde reaction capacity of DSP increased gradually from 145.4 mg/g to 243.1 mg/g with the degrading time increased gradually from 2h to 3.5h, indicating that the longer degrading time resulted in sufficient unfolding and degradation of soybean protein chains, very low viscosity at solid protein content of 37.5wt%. The new amino groups generated by amide hydrolysis are ready to react with aldehyde; and the higher formaldehyde reactivity indicated that the smaller degraded DSP chains were easily crosslinked by formaldehyde, glyoxal or reactive resin (such as UF and MF resin) to form polymer with larger molecular weight, or even the crosslinked product with good water resistance. It is apparent that the secondary and tertiary structures of globular soybean proteins were destructed during strong-alkali degradation, consequently, the polar groups that were wrapped on the globular structures of protein were released and were able to adsorb to polar wood surface to yield better bonding performance.Tab.2 The properties of degraded soy proteins with various timesArea percentage of each GPC peak / % Degradedtimes /h M1 M2 M3 M4 M5Formaldehyde reactivity /mg·g -1Viscosity /mPa·s (25o C) pH value 2 50.5 9.0 7.2 9.2 24.1 145.4 80.5 12.80 2.5 48.3 9.4 7.9 9.7 24.8 165.8 62.512.69 3 46.5 10.5 8.0 9.6 25.4 186.0 59.012.64 3.5 43.9 12.0 8.3 9.7 26.2 243.132.5 12.43Tab.3 The curing characteristics and bond performance of DSP based adhesivesDSC peak temperature at various heating rate /o C Bond strength /MPa Adhesive ID 15 10 7.5 5 2.5E a /kJ·mol -1Dry state Wet state J-AN/A N/A N/A N/A N/A N/A 0.63 N/A J-B152.1 143.5 139.0 133.1 124.0 83.6 0.73 N/A J-C179.6 176.0 168.6 160.3 150.4 84.2 1.10 N/A J-D189.6 181.5 178.1 169.8 159.5 91.4 1.03 N/A J-E212.3 205.3 198.7 187.6 183.2 93.7 1.26 0.64 J-F 204.7 194.4 190.6 184.3 173.6 96.8 1.25 0.70J-G 179.6 163.9 159.1 153.4 138.7 60 1.03 0.52J-H 182 170.7 167.2 154.9 137.6 53.4 0.87 N/A resin 100.4 93.1 89.8 83.8 76.9 76.8 1.18 N/A UFresin 180.5 178.2 174.7 169.9 160 113.4 1.62 1.21 PFFive DSP adhesives were prepared by blending the DSP (degraded for 3.5h) with glyoxal, UF and MF resin, respectively, as shown in Table 1. When the DSP solution was used alone as wood adhesive (J-A), DSC scanning did not detect any endothermic or exothermic peak, as shown by J-A curve in Figure 2, which indicated that there were no obvious chemical reactions as the system was heated. As for adhesive J-B that contained 23wt% of bifunctional glyoxal and 77wt% of DSP, the significant DSC exothermic peak at 124-152o C indicated that DSP could be crosslinked by glyoxal via reaction of aldehyde group with amino groups in DSP. Because the UF resin and MF resin contained hydroxyl methyl groups that could also react with some functional groups of soybean protein such as amino and amide group, the adhesives from J-C to J-F yielded DSC exothermic peaks similar to adhesive J-B during DSC scanning. The curing DSC peaks of UF resin and MF resin at different heating rates ranged from 76o C and 105o C. However, the adhesives prepared by blending DSP with UF or MF showed DSC curing peaks higher than 150o C and the absence of curing peaks of pure MF or UF resin below 105o C, indicating that the main curing reactions were underwent between DSP-UF or DSP-MF rather than UF-UF or MF-MF. The activation energy analysis in Table 3 showed that the blend of DSP with glyoxal (J-B) had the lowest activation energy, 83.6kJ/mol, and the blend of DSP with MF resin in the presence of glyoxal (J-F) had the highest value, 96.8kJ/mol. In the DSP-UF and DSP-MF blends, the presences of glyoxal led to higher activation energy of curing reaction, i.e., E J-D > E J-C and E J-F > E J-E, implying that the crosslinking reaction between DSP and synthetic resins were more difficult than that between DSP and glyoxal.Fig.3 DSC curves of adhesive J-G, J-H and PFR with heating rate at 2.5o C/minBecause the DSP solution with pH adjustment was strongly alkaline which adapted to the curing reaction of thermosetting PF resin, the following works focused on the new adhesive systems by blending DSP with PF resin or PF oligomer. Adhesive J-G was prepared by blending DSP and PF resin. Compared with bond strength of Adhesive J-A (DSP only), the addition of21.9% PF resin into DSP(liquid basis) increased the dry bond strength as expected, however, it did not yield acceptable water resistance for its bonded specimens had 0.52MPa of wet strength after 4h water boiling. Adhesive J-H was prepared by blending DSP and PF oligomers that mainly composed of multi-hydromethylated phenol. The oligomers could copolymerize with active groups of DSP and generated crosslinked macromolecular structure when heated up. However, introducing 27.8% of PF oligomer into DSP solution (liquid basis) did not yield good bond strength and water resistance because all wet-strength specimens were delaminated after 4h water boiling and the dry bond strengths were lower than that of J-G. This might be attributed to the lower corsslinking reactivity between PF oligomers and DSP and the lower cohesive strength of low-molecular-weight PF oligomers, consequently, the modified DSP did not form enough crosslinking density to resist boiling water. The bond strength and water resistance of adhesives J-G and J-H were much worse than that of PF resin only, as shown in Table 3. PF oligomers were active for they were ready to be crosslinked to form PF resin and then gelled when stored at ambient for days; the DSC scanning in Figure 3 confirmed that PF oligomers were more reactive than PF resin in terms of much lower curing peaking temperature (119.4o C vs. 160.5o C) and much larger exothermic peak. However, DSC scanning also indicated that the curing peak temperature of DSP-PF oligomer system (138.7o C) was comparable to that of DSP-PF resin system (137.6o C). Compared the DSC of adhesive J-G (DSP-PF resin) with that of PF resin and that of Adhesive J-H (DSP-PF oligomers) with that of PF resin, the curing peaks correlated to the PF resin (160.5o C) or PF oligomers (119.4o C) did not detected in their DSP blends (J-G and J-H), respectively. However, both systems were detected a new curing DSC peak at about 138o C which should be attributed to the crosslinking reactions of hydromethyl groups of PF resin or oligomers with amino groups of DSP. This fact also indicated that both PF resin and PF oligomers added were diluted by DSP so that their own curing peaks could not be detected at all; in other words, the reaction rate of DSP-PF resin (or DSP PF oligomers) was much faster than that of intermolecular PF resins (or PF oligomers).All DSP based adhesives except the J-A could be used to produce common indoor-used plywood that requires dry bond strength more than 0.7MPa according to Chinese standard GB/T 9846.3-2004. Of these adhesives, adhesive J-E and J-F (blends of DSP with MF resin) had the best dry strength (J-E: 1.25MPa; J-F: 1.26MPa); followed were the blends of DSP with UF resin (J-C: 1.03MPa; J-D: 1.10MPa) and the blend of DSP with PF (J-G: 1.03MPa) that were comparable to UF resin. Only J-E and J-F passed the 4h boiling-water test showed the potential to be used as water resistant adhesive; however they can neither pass the 28h boiling-dry-boiling test, indicating that they could not be used as outdoor weather-proof adhesive.3 ConclusionsBy degrading soybean proteins at 90o C in the presence of strong alkali, the advanced structures of soybean proteins were completely destroyed and polypeptide chains broke and, therefore, obtained DSP solution with molecular weight ranged from 282 to 3404, high protein concentration of 37.5wt% and very low viscosity. With the degrading time increased, DSP solution had less high-molecular components but lower viscosity and much more active groups in terms of much higher formaldehyde reactivity. The DSP could be crosslinked by glyoxal, UF resin, MF resin, PF resin and PF oligomers. The DSP based adhesives by blending DSP with glyoxal, UF resin, MF resin, PF resin and PF oligomer, respectively, could be used to produce common indoor-used plywood that requires dry bond strength more than 0.7MPa according to Chinesestandard GB/T 9846.3-2004, but only plywood bonded by MF/DSP blend tolerated 4h water boiling, showing the potential for using as water resistant adhesive.REFERENCES：[1] Hymowitz T, Collins F, Panczner J.Relationship Between the Content of Oil, Protein, andSugar in Soybean Seed. Agronomy Journal, 1972, 64(5): 613-616[2] Kumara R, Choudhary V, Mishra S. Adhesives and plastics based on soy protein products.Industrial Crops and Products, 2002, 16(3): 155–172[3] van der Leeden M, Rutten A, Frens G. How to develop globular proteins into adhesives.Journal of Biotechnology, 2000, 79(3): 211–221[4] Hettiarachchy N.S., KalaPathy U., Myers D.J. Alkali-modified soy protein with improvedadhesive and hydrophobic properties. 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J Am Oil Chem Soc, 2000, 77: 705–708 [11] Sun X, Ke B. Shear strength and water resistance of modified soy protein adhesives. Journalof the American Oil Chemists Society, 1999, 76( 8): 977 - 980[12] Gao Z, Wang M, Wan H, Liu Y. Curing characteristics of urea-formaldehyde resin in thepresence of various amounts of wood extracts and catalysts. Journal of Applied PolymerScience, 2008, 107(3): 1555 – 1562[13] Kalapathy U, Hettiarachchy N, Myers D. Alkali-Modified Soy Proteins: Effect of Salts andDisulfide Bond Cleavage on Adhesion and Viscosity. Journal of the American Oil Chemists Society, 1996, 73(8): 1063-1066通讯作者：高振华E-mail: gaozh1976@。
