Abstract

The environmental concerns about the formaldehyde emission from commonly used wood adhesives have put great attention to find a bio-based renewable substitute for them. Soy protein is abundant and inexpensive with great amount of polar functional groups such as hydroxyl, amino and carboxylic acid groups. These functional groups can form hydrogen bonds with hydroxyl groups of wood components, thus resulting in superior dry strengths of the resulting wood-based composite panels. However, the water resistant of soy based adhesives is very weak as water can easily disrupt these hydrogen bonds. Crosslinking the functional groups of SP with various curing agents is an effective strategy for improving the water resistance of the resulting wood-based composite panels. In this paper, various soy products, SP structure and characteristics, and different modification methods of SP was reviewed. Then, all the crosslinking agents developed along with their curing mechanism were deeply studied.

Keywords: crosslinking agent; soy protein; plywood; soybean flour; water resistance.

1.    Introduction:

The volume of the global wood adhesives and binders market was over 16,200 kilo-tons and it was valued at $13.15 billion at 2013 [1]. It is expected that the value of this market to reach $18.99 billion by 2022 [2]. Most of the wood adhesives are formaldehyde based and releasing formaldehyde during the production and use [3]. Formaldehyde was reclassified as a human carcinogen by the International Agency for Research on Cancer in 2004 [4]. The California Air Resources Board (CARB) passed a regulation on limiting formaldehyde emission from wood-based products used and sold in California in April 2007 [5]. A national regulation of limiting formaldehyde emission, ‘‘formaldehyde standards for composite wood products act,’’ was signed into law on July 7, 2010 [6]. Other adhesives highly used in the wood industry are isocyanate based resins, mostly in the form of Methylene diphenyl diisocyanate (MDI) and its polymer pMDI. These resins are very toxic and there are many health hazards associated with their usage during the production of panels [7]. Also, all these adhesives are petrochemical materials and the usage of them is not sustainable. Therefore, finding some renewable bio-based replacements for them is very crucial.

Many attempts were performed to produce wood adhesives from tannin, lignin, carbohydrate, unsaturated oils, and liquefied wood [8]. Proteins are another bio-based resource used for making adhesive. Animal based proteins like blood [9], collusion, and casein [10], and plant based proteins such as cottonseed [11], canola [12], wheat [13], peanut [14], corn [15], sorghum [16] were previously used for producing wood adhesive. However, most of the researches for manufacturing of wood adhesive focused on soy protein. Soy protein based adhesives were first developed in 1923 when a patent was granted for soybean meal-based glue [17]. The better properties of petroleum-based adhesive along with their cheap price were disastrous for the usage of soy in wood adhesive industry. It was only after the lapse of almost 70 years that the significance of ecofriendly adhesives has been realized and scientists begin to develop soy based adhesives again.

Soybean, originated from eastern Asia, is one of the most important crops in the US. Soybean oil and soybean meal are two major products of soybean. At present, soy bean meal is mainly used as animal feed. In the US, most soybean meal is consumed by livestock and poultry. Only a small portion of soybean meal is currently used in non-food industrial applications such as surfactants, inks, fuels, lubricants, and adhesives [18]. Soybean meal is abundant and inexpensive and mainly consists of soy protein and carbohydrates contains a great amount of polar functional groups such as hydroxyl, amino and carboxylic acid groups [19]. The polar functional groups of SF can form strong hydrogen bonds with hydroxyl groups of wood components, thus resulting in superior dry strengths of the resulting wood-based composite panels. However, water can easily disrupt the hydrogen bonds, thus dramatically reducing the water resistance of the resulting panels [20].

Reactions of the poplar functional groups with various curing agents for the reduction of the amount of the polar functional groups and the subsequent crosslinking of the SF have been found to be an effective strategy for improving the water resistance of the resulting wood-based composite panels [20]. In this study, a review was performed on various curing agents developed to convert soy protein into an insoluble water resistant adhesive for making wood composite panels.

2.    Soy Protein

Soy protein is composed of 18 different amino acids connected with each other through peptide bonds. The weight ratio and the number of moles of these amino acids in 100 g SP are shown in table 1. Each of these amino acids has a side chain with a specific functionality. Carboxylic acid, hydroxyl and amino groups are the most abundant side functional groups in SP. There are respectively 0.24, 0.11, and 0.12 moles of reactive carboxylic acid, amino, and hydroxyl groups in the side chains of 100 g SP. SP also contains a small amount of thiol functional group (0.01 moles in 100 g SP). The rest of the functional groups of SP are water hydrophobic and do not have any reactive side chain. These hydrophobic functional groups are folded inside the protein chain and make a globulin structure. Therefore, most of the active groups of SP are buried inside the globulin structures and are not accessible.

SP globulins interact with each other and form different quaternary structures. These structures consist of four major fractions, 11S, 7S, 2S and 15S, where the S is in Svedberg units as determined by sedimentation chromatography [21]. The 11S (glycinin) and 7S (β-conglycinin) are the most abundant fractions of SP and constitute 65–80% of the total the protein content [22]. Glycinin is a hexamer of six monomeric subunits (each comprises of two globulins) which are linked by single sulfide bonds. These subunits associated into two hexagonal rings forming a hollow cylinder held together by electrostatic and hydrogen interactions. Glycinin (11S) is dissociates into the 7S (six monomeric subunits) and/or the 3S (one monomeric subunits) forms. These dissociations are mainly driven by electrostatic repulsion depending on the solubility, pH and ionic strength. Glycinin features can also be associated into the 15S form containing twelve monomeric subunits [23, 24]. The range in molecular weight and approximate distribution of these fractions are as follows: 2S, 8–50 kDa, 8%; 7S, 100–180 kDa, 35%; 11S, 300–350 kDa, 52%; and 15S, 600–700 kDa, 5% [21].

SP are existed in different features, including defatted soy flour (SF), soy protein concentrated (SPC) and soy protein isolated (SPI). The main difference between these forms is their protein content which is approximately 50%, 70%, and 90% in SF, SPC, and SPI, respectively. There is a considerable amount of carbohydrates in the structure of SF and SPC. These carbohydrates are separated during the isolation process of SPI. Also, there are small amounts of ash and residue fat in all features [25]. The composition of SF, SPC, and SPI are shown in Table 2 [25-27]

Soy carbohydrates are the second-largest components in SF and SPC. Half of them in SF are non-structural carbohydrates like simple sugars and oligosaccharides along with structural carbohydrates such as cellulose, hemicellulose, pectin and starch [28, 29]. During the production of SPC, the soluble non-structural carbohydrates are washed and only insoluble structural carbohydrates are left [25]. All soy carbohydrates are highly hydrophilic and decrease the water resistance of SF and SPC adhesives. Also, most of them are not reactive enough and usually do not participate in any reaction. Steric hindrance and/or the high viscosity resulted from their solubility or swelling in aqueous solution also prevent the reaction between curing agents with SP functional groups, thereby reducing the efficiency of curing agents [28, 30]. Since all carbohydrates are removed in SPI, it has much more potential than SF and SPC to make a water resistant adhesive. However, the higher cost of SPI did not allow it to be used industrially for making adhesive yet.

Typically, to produce water resistant adhesive, it is necessary to crosslink the functional groups of SP. However, most of the functional groups of SP are buried inside its globular structures and are not accessible to react with crosslinking agent. Therefore, it is necessary to denature the structure of SP to expose its functional groups before adding any crosslinking agent. SP globular structures are sensitive to various conditions, such as pH, added denaturants and temperature [31]. Therefore, many different methods were developed for denaturing SP structure through applying heat [32, 33], enzymes [34, 35] and chemicals. The commonly used chemicals for denaturing SP are alkalis including sodium hydroxide [13, 33, 36, 37], urea [33, 37-40], and calcium hydroxide [41]; salts like guanidine hydrochloride [38, 42, 43], sodium bisulfite [44-46], and sodium sulfite [47]; and acids such as hydrochloric acid-, citric acid [48], and boric acid [48]. Also, different cationic detergents like hexadecyltrimethyl ammonium bromide, ethylhexadecyldimethyl ammonium bromide (EDAB), and benzyldimethylhexadecyl ammonium chloride [49] as well as anionic detergents like sodium dodecyl sulfate [37, 50, 51] and sodium dodecyl benzene sulphonate [52] were successfully used to denature SP. The denaturing effects of the combination of acid, salt, and alkali were also investigated [53]. Adjusting the pH value to isoelectric point of protein is another method used for denaturing SP without adding any chemicals [54]. Among different denaturing agents, the usage of sodium hydroxide, sodium dodecyl sulfate, and sodium bisulfite are the most effective methods commonly used.

3.    Crosslinking agents of SP
   1. Curing of SP with azetidium group
      1.     Polyamidoamine-epichlorohydrin resin (PAE)

PEA is a polymer highly used as a wet-strength agent for paper manufacturing [55, 56]. The results of the curing of SP via this polymer revealed that PEA resin is a great crosslinking agent for SP. The water resistance and the mechanical tests of the wood panels bonded with PAE-SP adhesive revealed that panels bonded with SPI-PAE and SF-PAE adhesives can be successfully used in exterior and interior applications, respectively [57-60]. A patented technology based on soybean flour and PAE resin has been successfully used in the commercial production of plywood and particleboard [58, 59, 61, 62]. PAE structure and its reactions to crosslink the SP are shown in Figure 1.

The azetidium groups in the PAE resin react with the secondary amines and carboxylic acid groups in the PAE structure, thus causing homo-crosslinking (reaction 1 and 2 in Figure 1-b). Also, the azetidium groups react with amino and carboxylic acid groups in SP structure (reaction 3 and 4 in Figure 1-b). All these reactions result in a water resistant 3-D network which is water resistant [57, 58, 63, 64].

The study of the viscosity and the bond strength between wood and the SF/PAE adhesive revealed that the viscosity of the adhesive was highly depended on the PDI and particle sizes of SF. However, neither of them had considerable effect on the bond strength between wood and the adhesive [65]. In another study, the bonding strength of SF/PAE adhesive at high temperatures was studied. The results revealed the decrease in the mechanical properties of plywood manufactured with SF/PAE adhesive is similar to those of solid wood, confirming the stability of SF/PAE adhesive at high temperatures [66].

The synthesis of PAE is typically a two-step process [67]. A polyamidoamine is first prepared, particularly by the polycondensation reaction of diethylenetriamine and adipic acid. The resultant polyamidoamine is then reacted with epichlorohydrin in an aqueous solution to form PAE solution. To synthesis PAE from renewable materials, some attempts were performed to either derive adipic acid from renewable materials or substitute it with citric acid or itaconic acid produced from renewable materials [67-70].

2.     Epichlorohydrin (ECH) and ammonium hydroxide (NH₄OH) reaction

Huang et.al. produced three curing agents named I, II, and III from the reaction of ECH and NH₄OH. Their chemical structures are shown in Figure 2 [71]. Compound I was prepared through the reaction of ECH and NH₃ in isopropanol under basis conditions. Compound I was directly reacted with NH₄OH without further purification to form II. Treatment of II with ECH in water resulted in III [71-74]. All the three curing agents were mixed with SF to produce adhesive for bonding wood.

The intermediates of the curing of SP by curing agent I are proposed in Figure 3. The same intermediates and mechanism are proposed for curing agent III. The OH group in the chlorohydrins (i.e., the 3-chloro-2-hydroxypropyl group) has a neighboring-group participation effect facilitating the formation of an azetidinium ring, an epoxy ring, or direct replacement of the chloride group with a nucleophilic amino group as it was shown in Figure 3 as (a), (b), and (c), respectively [75]. The azetidinium groups react with carboxylic acid and amino groups in the SP structure [57]. The curing agent I has three chlorohydrin groups and each of them might be converted to an azetidinium ring on heating under basic conditions. The chlorohydrin group might also be converted to an epoxy ring in alkaline solution [72, 76]. The generated epoxy ring could react with carboxylic acid or amino groups in SF, thus resulting in a crosslinked water-insoluble network. The direct replacement of the chloride group on the chlorohydrin groups with nucleophilic amino groups in SP is also possible. These amino groups could also lead to a crosslinking by forming salt bridges with the carboxylic acid groups of SP. The dehydration of the salts can lead to formation of amide linkages [71, 73, 74].

In the structure of curing agent II, all the chloride atoms of I were substituted with amino groups. They can form salts with carboxylic acid in SP and the dehydration of the salts can lead to formation of amide linkages. However, the effective formation of amides requires a relatively high temperature and long reaction time [77, 78]. Sufficient amounts of amides were unlikely formed from these salts under hot press temperatures and times used for making wood composites. Therefore, crosslinking in the SF-II adhesive mainly relied on the salt bridges between amino groups in II and carboxylic acid groups in SF. The salt bridges could be disrupted during the water soak test, which explains the failure of these specimens. Although II is not passing the industrial water resistant need of the plywood, the modulus of rupture, modulus of elasticity, and internal bond strength of particleboard panels bonded with the SF-II adhesive all exceeded the corresponding minimum industrial requirements for M-2 grade particleboard [71].

2. Curing of SP with oxirane ring

Oxirane ring can react with a wide variety of functional groups, such as hydroxyl, amino, mercapto, and carboxylic acid groups in SP [79]. Therefore, the polyepoxides are proposed to serve as crosslinking agents that convert flowable SP into water-insoluble crosslinked networks during the hot-pressing step of wood composite production. The reaction of an epoxy ring occurs easily in the case of the amino-glycidyl. It needs high temperature (200°C) to make hydroxyl-glycidyl and carboxylic acid-glycidyl reactions. When polyepoxide is curing SP, hydroxyl groups usually accelerate an amino-glycidyl reaction instead of reacting directly with glycidyl groups [80-82]. In the case of carboxyl acid groups, the dissociated form of COOH (COO−), which occurs in the presence of amines and hydroxides, reacts with the glycidyl group at 105°C. Therefore, aside from denaturing the structure of SF and exposing more functional groups for curing reaction, amines and hydroxides also dissociates carboxylic acid groups of SF to the form of COO− reacting with epoxy rings at much lower temperatures [19, 80-82].

1.     Triglycidylamine (TGA), glycerol polyglycidyl ether (GPE), and trimethylolpropane triglycidyl ether (TTE)

The plywood panels bonded with SF-TGA and SF-GPE adhesives did meet and those with SF-TTE adhesive did not meet the water-resistance requirement for interior application. The chemical structure of TGA, GPE, and TTE are shown in Figure 4-a. TGA (I in Figure 4-a) and TTE (IV in Figure 4-a) have very similar chemical structures, each containing three epoxy rings. GPE is a mixture that mainly consists of glycerol diglycidyl ether (II in Figure 4-a) and glycerol triglycidyl ether (III in Figure 4-a) with two and three epoxy rings, respectively [83, 84].

The proposed curing mechanism of SP via polyepoxides (V in Figure 4-b) is show in Figure 4-b. To be an effective crosslinking agent, a polyepoxide must be water soluble to be mixed well with SF in an aqueous solution. In the case of low water solubility, a polyepoxide tends to aggregate and thus the crosslinking will be poor. Plywood panels manufactured with SF-TGA and SF-GPE adhesives were water resistant and the reason is the good solubility of TGA and GPE in water. Also, the tertiary amino group existed in the structure of TGA is known as catalyst for activating the epoxy resins [84], thus making TGA even the better crosslinking than GPE. Panels bonded with SF-TTE failed the required water resistant test and the reason is poor water solubility of TTE [85].

2.     Poly (glycidyl methacrylate-co-styrene) (PGS)

PGS was synthesized through an emulsion polymerization of glycidyl methacrylate and styrene monomers. PGS is a stable emulsion that contains small and well-dispersed PGS chains in water, and can mix and react well with the polar functional groups of the SF components for the crosslinking reactions [20, 86]. The crosslinking mechanism of SP via PGS is shown in Figure 5.

The results revealed that PGS is a great curing agent for SP if it is incorporated with at least 2.5 wt% of NaOH. Since PGS is a polymer with long chains, it cannot easily diffuse into the structure of SP. Therefore, a high amount of NaOH is necessary to denature the structure of SP and expose its functional groups to react with PGS. Also, in the presence of NaOH, the carboxyl acid groups are dissociated to the form of COO⁻ reacting with the glycidyl group at 105°C (rather than 200°C)[19, 20, 80-82].

In another attempt to use PGS as the curing agent of SP, an adhesive was produced by mixing PGS, polyethyleneimide (PEI) and SF [87]. The crosslinking mechanism of this adhesive is shown in Figure 6.

When PGS and SF are mixed first, the epoxy rings of PGS react with some amino groups of SP at room temperature. However, most of the functional groups of SP are carboxylic acid rather than amino groups and many of them are not accessible for PGS. Also, there is not enough time for completing the reaction of glycidyl and amino groups. The possibility of chemical reaction between oxirane rings of PGS and carboxylic acid groups of SP is not so high since it needs high temperatures. Therefore, a lot of oxirane rings remain unreacted. These unreacted epoxy rings chemically react with the amino groups of PEI when it is added later. Therefore, PGS chains act as bridges to connect the SP and PEI structures and to make the whole system crosslinked. Also, formation of a few amide linkage through the reaction of amino groups of PEI and carboxylic acid groups of SP during the hot press is possible [87].

3.     1,3-dichloro-2-propanol (DCP)

The curing mechanism of SP via DCP is shown in Figure 7.

With catalysis by NaOH, DCP would likely first form epichlorohydrin (1 in Figure 7) that could further react with many nucleophiles, such as -NH₂, -OH, -SH, and -COOH groups in SP. For instance, the reaction of epichlorohydrin (I in Figure 7) with amino and carboxylic acid groups in SP would yield 2 and 5, respectively. The newly formed 2 and 5 could be further converted to epoxide-containing SP 3 and 6, respectively. Reaction of 3 with amino and carboxylic acid groups in SP would result in 4 and 8, respectively. Reaction of 6 with -NH, and -OH groups in SP would afford 7 and 8, respectively. The epoxide group in 1, 3, and 6 could also react with -SH and -COOH groups in SP [88].

4.     Ethylene glycol diglycidyl ether (EGDE) and diethylenetriamine (DETA)

SF was also cured via a mixture of EGDE and DETA. DETA reacted with EGDE to form a network structure containing a lot of active epoxy groups (Figure 8). Just like the usage of PGS for curing of SF (part), since the active epoxy rings are connected to the EGDE/DETA network, they cannot diffuse into the SP structure to react with its functional groups. Therefore, denaturing of SP to expose its functional groups before making reaction with the active epoxy rings of the EGDE/DETA network is so necessary [89].

In addition to the chemical reaction of epoxy rings of EGDE/DETA network with the functional groups of SP, the EGDE/DETA network formed an interpenetrating network with the SP molecules, which further improved the water resistance of the adhesive [89]. A schematic view of these interpenetrating networks is shown in Figure 9.

5.     Bisphenol A diglycidyl ether (BPADGE) resin

To make a water resistant adhesive, the usage of a BPADGE resin as the curing agent of SP was studied as well. BPADGE resin cannot be dissolved into water, so using a surfactant to disperse BPADGE chains is necessary. Each nearly long chain of the BPADGE has only two active epoxy groups which can react with the functional groups of SP. Therefore, in order to make enough crosslinking, adding high amounts of BPADGE in necessary. When the weight percentage of BPADGE was more than 30% of the total solid adhesive, the plywood panels might pass the required water resistant test. At this high concentration, BPADGE molecules tended to aggregate and also increase the viscosity of the adhesive. Because of these problems, BPADGE resin was rarely used as the curing agent of SP alone, and it was usually used along with other curing agents [90-92].

3. Curing of SP with metal components
   1.     Magnesium oxide (MgO)

The mechanisms by which MgO interacts with soy components are proposed in Figure 10. MgO particles have rigid, porous structures and are widely used as a desiccant for books and foods [93-95]. They tend to absorb water on their surfaces without destruction of the MgO structures [96-100]. When MgO particles are mixed with SF in water, the soy components (carbohydrates and proteins) would get into the gaps and voids of MgO particles. Soy components contain numerous hydroxyl and amino groups that can form hydrogen bonding with MgO. Soy proteins also contain high amounts of carboxylic acid groups that can react with MgO to form a magnesium carboxylate salt. The carboxylic acid groups are spread along the soy protein chains. The chance that two carboxylic acid groups on the protein chains would react with the same Mg²⁺ atom to form a –COOMgOOC– salt should be rare. Therefore, the magnesium carboxylate salt is highly likely to be a half salt (–COO–Mg–O–MgO). A heat treatment may facilitate the reactions between the carboxylic acid groups and the surface MgO and the removal of the resulting water from the reactions, thus resulting in a SP–MgO complex where most of the carboxylic acid groups are buried inside MgO. Once the complexes were formed, they became very water resistant because it was hard for water to penetrate and disintegrate the complexes [101, 102].

2.     Nano calcium carbonate crystalline arrays

SP/nano-CaCO₃ adhesive was prepared by co-precipitating calcium carbonate in SPI alkaline aqueous solution. Inspired by gecko (Figure 11-c) and mussel adhesion (Figure 11-g), the biomimetic SP/nano-CaCO₃ adhesive showed good adhesion effects on wood adherents (Figure 11-e) even after water soaking and drying cycles because of interlocking forces and ionic crosslinking . Wettability, adhesion strength, and water-resistance of the SP/nano-CaCO₃ glue showed great advantages over pure SP adhesive. Firstly because calcite of CaCO₃ with nano or sub-micron-scaled arrays along the surface of SP could easily penetrate into wood lumens to form the compact rivets or interlocking links. Secondly, calcium, carbonate, and hydroxyl ions acted as ionic crosslinkers to bind SP chains together and to wood. Ionic bonding exhibits a much higher strength and resistance to water than hydrogen bonding. The plywood samples manufactured with this adhesive showed stable adhesion strength more than 6 MPa even after water soaking and drying cycles, indicating that SP/nano-CaCO₃ presented either a good internal bonding strength between protein and CaCO₃ or good external glue strength between adhesive and wood substrate [103, 104].

3.     Magnesium aluminium phyllosilicate (attapulgite)

Attapulgite is a silicate clay mineral, which has a layered chain structure containing water rich magnesium aluminium. Attapulgite has a special rod-like fiber structure and a great deal of hydroxyl groups on its surface. The chemical and the rod-like fiber structure of attapulgite are shown in Figure 12. The hydroxyl groups on the structure of attapulgite make hydrogen bonds with the functional groups of SP. Also, the magnesium ions of attapulgite form salt with the carboxylic acid groups of SP. The rod-like structure of attapulgite might fill the holes and cracks of the adhesive and act as a bridge joint between SP chains and make it physically and ionically crosslinked [105].

4.     Organic calcium silicate hydrate (CSH)

The proposed mechanism leading to a water resistant adhesive from SP/CSH is shown in Figure 13. Initially, the two silicate precursors, which are 3-aminopropyltriethoxysilane (APTES) and tetraethoxysilane (TEOS), are hydrolyzed under acidic conditions, leading to silanol formation (reactions 1 in Figure 13). The silanols conduct polycondensation reaction and connected with each other via very stable siloxane bonds. There are some evidences that silanol groups also might conduct polycondensation reaction with the hydroxyl groups of wood and SP as well [106]. Also, the hydroxyl groups of silanols can be ionically crosslinked around Ca⁺² ions to form the CSH hybrids [107, 108]. APTES is a molecule that carries two different reactive groups on its silicone atom and as shown in reaction 3 of Figure 13. Amino group of APTES could react with carboxylic acid groups of SP to form amide linkage in high temperatures. As a result, bridges which are connecting SP chains to the CSH phases are made making the whole mixture crosslinked.

There are some other studies conducted very similar experiments, but used active epoxy ring rather than amino group on the other end of the silicone group. As a result of the reaction between epoxy ring in the one end of the curing agent and the functional groups of SF, a similar crosslinked structure as it was shown in Figure 13 is formed that makes the wood composites water resistant [106].

4. Curing of SP through Hydroamination reaction
   1.     Maleic anhydride (MA) and polyethyleneimine (PEI)

For making this adhesive, PEI and MA were first mixed in an alkaline condition. As a result, MA reacted with PEI to form amide-linked maleyl group. Then, SF was added to the mixture of PEI and MA and after mixing, the adhesive was spread onto veneer. During a hot-pressing of panels, the amino groups in SF and PEI can react with carbon-carbon double bong of the MA as it is shown in Figure 14. As a result, MA groups act as bridges to connect the PEI and SP structures and to make the whole system crosslinked [18, 85, 109-112].

Because PEI is expensive, some researches were performed to replace it with other polyamines. In one study, a new type of polyamidoamine (PADA) resin was synthesized by the polycondensation reaction of polyethylene polyamine and adipic acid. This synthesized polyamidoamine was substituted PEI and was successfully used with maleic anhydride (MA) and SF to make adhesive [113].

2.     Glutaraldehyde and polyglutaraldehyde

Aldehydes have a potential to be used as a crosslinking agent of proteins. Studies of collagen protein crosslinking reactions with monoaldehyde (formaldehyde) and dialdehydes with two to six carbon atoms (glyoxal, malonaldehyde, succinaldehyde, glutaraldehyde, and adipaldehyde) demonstrated that the reactivity is maximized at five carbons; thus glutaraldehyde is the most effective crosslinking agent [114].

Aldehydes are expected to form Schiff bases upon nucleophilic attack by the amino groups of protein. However, Schiff bases are unstable and tend to break down to regenerate the aldehyde and amine. In contrast, the linkage formed by the reaction of glutaraldehyde with an amino group has shown exceptional stability at extreme pHs and temperatures, thus a simple Schiff base with both ends of monomeric glutaraldehyde (reaction 1 in Figure 15.) has been ruled out as a mechanism for glutaraldehyde crosslinking with proteins [115].

Glutaraldehyde polymerize through a Schiff base spontaneously in aqueous solutions, at room temperature in the absence of any catalyst (reaction 2 in Figure 15.). Commercial solutions of glutaraldehyde are largely oligomers and polymers of glutaraldehyde and contained significant amounts of α,β-unsaturated bonds. A proposed curing reaction of protein via glutaraldehyde involved the conjugate addition of protein amino groups to double bonds (Michael-type addition shown in reaction 3 in Figure 15.). A different proposed mechanism involve an addition reaction occurred on the aldehydic part of the α,β-unsaturated polymers (reaction 4 in Figure 15.). Contrary to the Schiff bases with monomeric glutaraldehyde, Schiff bases with polymeric glutaraldehyde are stabilized via conjugated α,β-unsaturated bond through resonance formation. The exact curing mechanism of proteins with glutaraldehyde is not clearly understood yet. Some other mechanisms were proposed for the crosslinking reaction of proteins with glutaraldehyde in [115]. However, it seems that no single mechanism is responsible for glutaraldehyde reaction with proteins and several of these possible reaction mechanisms could proceed simultaneously [116-120].

Glycidyl methacrylate monomer (GMA)

The epoxy ring existed in one of ends of GMA monomer react with functional groups of SP, especially amino and carboxylic acids groups. The GMA monomer can react with the amino groups of SP with the carbon-carbon double bond through Michael-type addition reaction. As a result, GMA plays the role of a crosslinking agent to block the hydrophilic amino and carboxylic functional groups of SP chains and crosslink them [121].

5. Curing agents based on isocyanate functional group

Isocyanate functional group can react with most groups that possess active hydrogen atoms in proteins, such as OH, COOH, NH₂, NH, and SH (Figure 17) [122]. Therefore, SP can be crosslinked by polyisocyanate to form a network structure. Due to the high reactivity of the isocyanate group, the viscosity of the polyisocyanate-modified soybean protein adhesive increases rapidly and the crosslinking structure is not usually uniform throughout the adhesive. This results in a considerably short work life of approximately 30 min. However, the work life is one of the most important technical parameters of wood adhesives, because a sufficiently long work life not only provides sufficient working time for processing, but also ensures good bond properties by avoiding pre-curing prior to hot pressing. Therefore, the work life of polyisocyanate-modified adhesive should be improved for commercial applications [123-125].

In an attempt to extend the work life of SP/ polymeric Methylene Diphenyl Diisocyanate (pMDI) adhesive, nano-scale montmorillonite (MMT) platelets was used. The retarding mechanism of MMT platelets is shown in Figure 1. When SP and MMT were mixed, the structure of MMT was destroyed by the intercalations of SP chains. As a result, nano-scale MMT platelets were exfoliated into SP matrix. These MMT plates temporary blocked the functional groups of SP by forming hydrogen bonding with them. Also, they increased the steric hindrance between functional groups in SP and pMDI. Taking the illustration in Figure 18 as an example, the free SP molecules that do not interact with the exfoliated MMT platelets (labeled as SP-0) can freely react with pMDI, while those interacting with MMT platelets (labeled as DSP-1, -2, -3, -4 and -5) cannot. SP-1 has three active sites, while SP-2, -3 and -4 have one or two active sites available for bonding. SP 5 lacks free groups and cannot form any bond. Only 10 of the 18 active groups in the six SP molecules in the illustration in Fig. 8 are able to react with pMDI. Also, the un-bonded NH2 in SP-4 cannot react with pMDI due to the steric hindrance by the MMT platelets. As a result, the SP/pMDI/MMT adhesive exhibited a longer work life [123, 124].

Typically, decreasing the pMDI content leads to the longer pot life of SP/pMDI adhesive. However, little amounts of pMDI cannot produce enough crosslinking to make wood panels water resistant. Some researchers tried to use other curing agents beside pMDI to produce water resistant wood panels with lower amounts of pMDI. The simultaneous usage of 10 wt% of bisphenol-A epoxy resin along with only 6, 5, 4, 3, 2, and 1 wt% of pMDI as the co-curing agents of SP was considered a successful strategy for production of wood panels with satisfying properties [126]. In another attempt to make a soybean based adhesive with 70-80 wt% natural materials, aqueous solution of glyoxalated soy flour and condensed tannin was cued with pMDI. The resulted adhesive could be used in acceptable processing conditions and the manufactured wood composites shown to yield results satisfying the relevant standard specifications for interior wood boards [127]. Another researchers replaced 50 wt% of polyisocyanate needed to cure SP with glyoxal. Glyoxal contains two active aldehyde groups that are able to crosslink soybean proteins via Maillard type chemical reactions, but they are much less reactive than the isocyanate groups of pMDI. This combination could effectively balance the low reactivity of glyoxal and high reactivity of polyisocyanate, forming an even crosslinking structure during suitable time [125, 128].

6. Curing agents based on polyketones

[129]

7. Maillard reaction

Viscozyme L can effectively hydrolyze the polysaccharides existed in the SF and SPC to reducing sugars such as galactose, glucose, and arabinose. These sugars can cure SP through Maillard reaction, as it is shown in Figure 20 [130-132]. Since the produced monosaccharides are not so reactive to diffuse inside the structure of SP and react with its functional groups, denaturing the structure of SP to expose its functional groups can effectively improve the bonding strength and the water resistant of wood composite products [133-135]

In another attempt to cure SF without any external crosslinking agent, carbohydrates of SF were separated, hydrolyzed to monosaccharide, and oxidized to convent some of the hydroxyl and aldehydes groups to carboxylic acid functional group. Then, they were added again to SP to crosslink it through amide linkage formed by the reaction of carboxylic acid groups of the curing agent and the amino groups of SP and also through maillard reaction [136].

8.  Formaldehyde based resins

Some attempts were performed to crosslink SP with formaldehyde. However, none of them led to a stable 3-dimensional structure. Formaldehyde reacts with amino groups of SP through different mechanisms. It forms a Schiff base upon nucleophilic attack by amino groups of SP as it is shown in reaction 1 in Figure 21. However, since there is not any conjugated double bond to stabilize this Schiff base through resonance, this reaction easily moves back in the reverse direction by changes of pH and/or temperature. At 75°C to 90°C, it is also expected that formaldehyde react rapidly with chain-terminating amino groups of lysine, histidine, arginine, and tryptophan, two ortho positions of phenol in tyrosine, and thiol group in cysteine [137], as it is illustrated for a primary amino group in reaction 2 of Figure 21. The addition of formaldehyde stabilizes denatured protein chains, so they would not return to their native forms [137]. The amount of formaldehyde that is required to saturate SF is highly depended on the protein percentage. Also, the size of the particles of SF and the amount of the denaturation of protein highly determine the amount of the exposed functional groups of SP and therefore the amount of formaldehyde required to saturate them. Typically, It is in the range of 10-20 wt% of SF [138]. It is expected that these methylol groups sit on the functional groups of SP to react with the amino and methylol groups of other chains of SP. Therefore, a crosslinked structure should be made through methylene and ether bridges, very similar to the 3-D structure of formaldehyde based resins. A schematic of these reactions is shown in the reaction 2 and 3 of Figure 21. However, it was observed that none of these products is stable and they can easily move in the reverse direction, breaking the crosslinks. To stabilize these bonds, higher amounts of formaldehyde must be used which is not acceptable because of health issues. Therefore, formaldehyde-modified soybean flour is very water soluble, and thus extractable, even after oven drying at 150°C, suggesting it is not possible to successfully crosslink SP with formaldehyde alone [137].

The usage of formaldehyde based resins like phenol-, urea-, melamine-, resorcinol-formaldehyde resins or a combination of them had been very successful for crosslinking SF. The mechanisms of crosslinking of SP with PF as an example of formaldehyde based resins are shown in Figure 22. This mechanism is exactly the same as the polymerization and curing of phenol with formaldehyde, and the only difference is the participation of the functional groups of SP [139, 140]. The methylol groups in PF can react with two ortho positions of phenol group of tyrosine (reaction 1) and the amino groups of arginine (reaction 2), lysine (reaction 3), histidine (reaction 4), and tryptophan (reaction 5) in the SP structure. Also, the formation of the ether links through the reaction of methylol groups of PF with methylol groups resulted from the reaction of formaldehyde with SP (reaction 2 in Figure 21) is also possible to form less stable bonds. The usual polymerization of PF resin comprising of the condensation reactions of methylol groups with ortho and para positions of phenol and other methylol groups as shown in reaction 6 and 7 are still the main reactions leading to a 3-D network similar to Figure 23 [138]. The type and the density of the intra and inter crosslinking bonds between formaldehyde based resins and SP is highly depended on pH, leading to optimum adhesion strength at a specific pH depending on the resin type.

Typically, three-step processes for making durable soy-based adhesives with a formaldehyde based resin are: (1) denaturation of SP through different approaches, (2) modification of the denatured SP with formaldehyde and (3) co-polymerization of modified denatured SP (2) with a formaldehyde based resin [138]. Hse et al replaced 30 wt% of phenol with caustic hydrolyzed SF for making flakeboard panels. The final panels passed the exterior flakeboard panel requirements [141]. In another study, a soy based adhesive was used for producing random strandboards. The results revealed that the adhesive with 40 wt% SF was equal in performance to the PF when used under the same pressing conditions. Higher percentages of soybean can be used if longer press times are utilized. Also they showed that PF resin may be added by either in situ preparation or post-reaction blending. The final copolymer resins in both methods result in a water-durable adhesive [137, 142]. The effect of the PF viscosity on the bond quality of SF-PF adhesive with wood was studied as well. Higher viscosity of the PF resin resulted from the higher molecular weight of the PF chains increased the bond quality mainly by limiting the over penetration of the adhesive into wood structure. It is easy to understand that very high viscosity do not lead to a better bond quality as well [143]. In other studies, the performance of SF/MUF (maleic urea formaldehyde) was studies. The addition of MUF resin, not only significantly decrease the viscosity of SF-based adhesive but also increase its water-resistance and wet shear strength values. The results showed that this adhesive can to be used for producing interior plywood. Also, in all ratios of SF/MUF, the amount of formaldehyde emission was much less than the pure MUF [140, 144, 145].

Although the curing of SF with formaldehyde based resins do not completely solve the problem of formaldehyde emission from formaldehyde based resins, but it decreased the amount of the formaldehyde emission. For example, the formaldehyde emission of SF-PF adhesive was measured to be 0.32 mg/L completely meeting all the formaldehyde emission standards [146].

4.    Enhancing the water resistant of SP through grafting methods
   1. Grafting mercapto groups on SP

Inspired by the mussel protein which is a strong and water-resistant adhesive, the free –SH group was introduced into the SP by an amide linkage (Figure 24). First, acetylation of the –SH group in I yielded S-acetylcysteamine II. The reaction of the amino group in II and the free carboxylic acid groups in the SP in the presence of EDC, as catalyst, provided S-acetylcysteamine-modified SPI (III). The treatment of III with a NaOH solution removed the acetyl group to provide cysteamine-modified SPI (IV) [147].

Increasing the –SH content of SP dramatically increased the strength and the water-resistance of the wood composites. the –SH groups in a protein are easily oxidized to form disulfide bonds, thus cross-linking the protein to form a three dimensional network [148, 149]. The high temperature in the hot-pressing of the wood composites would certainly facilitate such a crosslinking reaction. The –SH group can also react with quinones, which is resulted from tyrosine, through a Michael addition reaction [150]. The pH value of the modified SP was about 10; it was highly likely that a part of the tyrosine moieties in the modified SPs oxidized to form quinones during the hot-press. Therefore, covalent linkages between the –SH group and the tyrosine moieties could not be ruled out [147].

2. Grafting dopamine (DA) group on SP

Mussel protein is a strong and water resistant adhesive. It contains high amounts of 3,4-dihydroxyphenylalanine (DOPA). Inspired by mussel protein, dopamine group was grafted on SP. Preparation procedure of dopamine-modified SP is shown in Figure 25. Protection of phenolic hydroxyl groups in 1 with dichlorodiphenylmethane readily yielded 2. When SP was treated with 2 in the presence of EDC, as the catalyst, the amino group in 2 reacted with the carboxylic acid groups in SP to form amide linkages (3). Deprotection of 3 readily provided 4 (SP-DA) [151].

Grafting of DA to SP greatly increased the shear strengths and water-resistances of plywood samples bonded with SPI-DA adhesive. The mechanism of this soy based adhesive is exactly the mechanisms by which marine mussel adhesives work. In the marine adhesive, the tyrosine residues are first hydroxylated by an enzyme to DOPA residues that are subsequently oxidized to quinones. Various inter- and intra-peptide crosslinkages among tyrosine/DOPA residues and between quinones and other amino acids such as lysine finally transform the marine adhesive protein into a tough and insoluble material. This crosslinking process is called quinone-tanning [152]. Also, the adjacent two phenolic hydroxyl groups in DOPA play very important roles in forming hydrogen bonds between adhesives and their substrates. The adhesion mechanisms for the SPI-DA adhesive are proposed to be similar to the natural quinone-tanning process. Because the two phenolic hydroxyl groups in DA or DOPA are easily oxidized to form quinones at an elevated temperature of hot press without the presence of any catalyst, the same crosslinking reactions happens in the SPI-DA adhesive [151].

3. Grafting undecylenic acid (UA) on SP

The carboxylic acid group of UA and the amino group of SP can carry out amide linkage in the present of EDC catalyst as Figure 26. EDC reacts with a carboxylic acid group of UA first and forms an intermediate reactive O-acylisourea that quickly reacts with an amino group of SP to form an amide bond [153, 154]. However, EDC didn’t catalyze the reaction between carboxylic acid and amino groups of protein chains. To catalyze protein self-crosslinking reaction, it needs other catalyst besides EDC and different conditions [154-156].

4.4. Phosphorylation of SF

Phosphorus oxychloride (POCl₃) reacts with amino and hydroxyl groups in proteins [157]. In addition to proteins, carbohydrates of SF such as cellulose, curdlan, and dextran can be phosphorylated using phosphorylating agents, predominantly at the C-6 hydroxyl group [158]. These reactions are shown in Figure 27. Thus, it can be envisioned that reaction of POCl₃ with SF would result in phosphorylation of both proteins and polysaccharides in SF [159].

Chemical phosphorylation of SF dramatically increased its wet bond strength. The attached phosphate groups acted as cross-linking agents, either via covalent esterification or ionic and hydrogen-bonding interactions with hydroxyl groups on protein and wood [159].

4.5. Grafting 2-octen-1-ylsuccinic anhydride (OSA) on SP

Liquid OSA, which have an oily nature and possesses a long hydrophobic alkyl chain, was used to modify SP. OSA was grafted onto SP molecules through a reaction between amino and hydroxyl groups of SP and anhydride groups of OSA [160, 161], as it is shown in Figure 28.

The wet adhesion strength of SPA was greatly improved by using up to 3.5% OSA modifications due to the oily nature and introduced hydrophobic long alkyl chains of OSA. However, this improvement leveled off as OSA concentration further increased beyond 3.5%, which are mainly attributed to the poor wetting ability of the modified SPA because of the excessively enhanced electrostatic and hydrophobic interactions [161].