We investigated the effects of ultrasound treatment on cellulase adsorption and lignocellulose hydrolysis. The activity of cellulase remained constant upon low-power ultrasound treatment (<120 W) and decreased using high-power ultrasound (>280 W). Oscillating cellulase adsorption occurred upon ultrasound treatment with any intensity. The maxima for desorption and adsorption were 41.9 and 83.1%, respectively, during 1 h of 90 W ultrasound treatment at 50 °C. A comparison between the short-time with long-time ultrasound experiments indicated that ultrasound treatment tended to desorb cellulase from substrate. However, ultrasound treatment also led to further surface erosion of biomass, which increased cellulase accessibility. These joint actions of ultrasound treatment induced the oscillating adsorption of cellulase. The increase in cellulase accessibility caused by ultrasound treatment led to a significant enhancement in lignocellulose hydrolysis.

In this study, we proposed a bioinspired approach for the deposition and zwitteration of hyaluronic acid (HA) with a reduced glutathione (GSH) to form a composite layer that functions as a low fouling coating. A polyanion of the HA–dopamine conjugate (HADA) possessing catechol groups was synthesized by carbodiimide chemistry between HA and dopamine. Then, the dopamine conjugated biofunctional polymers (HADA) were grafted onto Au substrates via the transformation of catechol into a quinone group under mild oxidative conditions followed by a reaction with GSH to avoid undesired adhesion and also to shield the exposed Au substrate. Analysis of XPS spectra and wettability indicated that HADA and GSH were successfully grafted onto Au substrates. Surface plasmon resonance analysis showed that both HADA and further GSH modified surfaces exhibited reduced nonspecific adsorption. The attachment of GSH to HADA modified surfaces (HADA-G) resulted in better antifouling performance, with a low or ultralow protein adsorption of 0–7.51 ng cm−2 when exposed to single protein solutions, and a reduction in nonspecific adsorption from cow's milk to 10% compared to that of bare gold. The enhanced antifouling performance of HADA-G modified surfaces was likely due to the zwitterionic structure in GSH, which can induce stronger surface hydration through electrostatic interactions as well as the hydrogen bonding induced by HADA. Our results provide a facile and universal approach to surface modification and demonstrate the benefits of using a composite layer for the design of low fouling surfaces.

This study presents a facile, rapid and effective method for the fabrication of optical fiber surface plasmon resonance (SPR) sensors via polydopamine (PDA)-accelerated electroless plating (ELP). The bioinspired PDA coating formed through the facile self-polymerization of dopamine (DA) was utilized as a versatile material for optic-fiber functionalization. Gold seeds were then rapidly and firmly adsorbed onto the PDA functional layer by amino and imino intermediates generated during the polymerization, and a gold film sensor was fabricated after metal deposition. The fabrication time of the sensor was decreased by 6–12 times, and the fabricated sensor exhibited higher sensitivity, better reproducibility and adhesion stability compared with those fabricated by the traditional ELP. Some key experimental parameters, including DA polymerization temperature, DA polymerization time, and plating time, were investigated in detail. The optimized sample exhibited high sensitivity ranging from 1391 nm per RIU to 5346 nm per RIU in the refractive index range of 1.328 to 1.386. Scanning electron microscopy images indicated that the sensor surface consisted of gold nanoparticles with a uniform particle size and an orderly arrangement, and the film thickness was approximately 60 nm. Another PDA layer was formed on the gold film for facile immobilization of antibodies. The sensor exhibited effective antibody immobilization ability and high sensitivity for human IgG detection over a wide range of concentrations from 0.5 to 40 μg mL−1, which indicate the potential applications of the fabricated sensor in immunoassays.

A catalytic membrane reactor, which contains a membrane matrix and a catalytic film of alloy nanoparticle-loaded β-lactoglobulin fibrils (NPs@β-LGF), was developed for the continuous-flow reduction of 4-nitrophenol (4-NP). The Cu–Ag and Cu–Ag–Au alloy NPs were synthesized using β-LGF as a scaffold and stabilizing agent. In this process, the Cu nanoclusters were formed in the initial stage and were able to promote the synthesis of Ag0, which acts as a reducing agent for the rapid formation of Au0. Furthermore, a catalytic membrane reactor was constructed by depositing the NPs@β-LGFs on a membrane matrix. The catalytic activity of the Cu–Ag–Au alloy NPs was higher than that of the Cu–Ag alloy NPs, using the reduction of 4-NP to 4-AP as a model reaction. The observed rate constant in the continuous-flow system is also higher than that in the batch system. In addition, these catalytic membrane reactors had good operating stability and antibacterial activity.

Bionanomaterials synthesized by bioinspired templating methods have emerged as a novel class of composite materials with varied applications in catalysis, detection, drug delivery, and biomedicine. In this study, two kinds of cross-linked lysozyme crystals (CLLCs) with different sizes were applied for the in situ growth of Au nanoparticles (AuNPs). The resulting composite materials were characterized by light microscopy, scanning electron microscopy, transmission electron microscopy, thermogravimetric analysis, and X-ray photoelectron spectroscopy. The catalytic properties of the prepared materials were examined in the catalytic reduction of 4-nitrophenol (4-NP) to 4-aminophenol (4-AP). It was found that the size of the AuNPs increased with an increase in Au loading for both small and large crystals. In addition, small crystals favored homogeneous adsorption and distribution of the metal precursors. And the size of the AuNPs within small crystals could be maintained below 2.5 nm by managing the HAuCl4/lysozyme molar ratio. Furthermore, the lysozyme functional groups blocked the AuNP activity sites, therefore reducing their catalytic activity. This effect was more pronounced for small AuNPs. Moreover, the mass transfer of reactants (4-NP) from solution to AuNPs within the crystals restricted their catalytic reduction, leading to superior catalytic performance of the AuNPs within small cross-linked lysozyme crystals (Au@S-CLLCs) compared to those within large cross-linked lysozyme crystals (Au@L-CLLCs) at similar Au loadings. Finally, an increase in Au loading clogged the crystal channels with increased quantities of larger AuNPs, thus impeding the catalytic performance of Au@S-CLLCs.

The aim of this study was to explore the influence of amphiphilic and zwitterionic structures on the resistance of protein adsorption to peptide self-assembled monolayers (SAMs) and gain insight into the associated antifouling mechanism. Two kinds of cysteine-terminated heptapeptides were studied. One peptide had alternating hydrophobic and hydrophilic residues with an amphiphilic sequence of CYSYSYS. The other peptide (CRERERE) was zwitterionic. Both peptides were covalently attached onto gold substrates via gold–thiol bond formation. Surface plasmon resonance analysis results showed that both peptide SAMs had ultralow or low protein adsorption amounts of 1.97–11.78 ng/cm2 in the presence of single proteins. The zwitterionic peptide showed relatively higher antifouling ability with single proteins and natural complex protein media. We performed molecular dynamics simulations to understand their respective antifouling behaviors. The results indicated that strong surface hydration of peptide SAMs contributes to fouling resistance by impeding interactions with proteins. Compared to the CYSYSYS peptide, more water molecules were predicted to form hydrogen-bonding interactions with the zwitterionic CRERERE peptide, which is in agreement with the antifouling test results. These findings reveal a clear relation between peptide structures and resistance to protein adsorption, facilitating the development of novel peptide-containing antifouling materials.Keywords: antifouling; biosensor; nonspecific adsorption; peptides; SPR;

A versatile, convenient, and cost-effective method that can be used for grafting antifouling materials onto different surfaces is highly desirable in many applications. Here, we report the one-step fabrication of antifouling surfaces via the polymerization of dopamine and the simultaneous deposition of anionic hyaluronic acid (HA) on Au substrates. The water contact angle of the Au surfaces decreased from 84.9° to 24.8° after the attachment of a highly uniform polydopamine (PDA)/HA hybrid film. The results of surface plasmon resonance analysis showed that the Au-PDA/HA surfaces adsorbed proteins from solutions of bovine serum albumin, lysozyme, β-lactoglobulin, fibrinogen, and soybean milk in ultralow or low amounts (4.8–31.7 ng/cm2). The hydrophilicity and good antifouling performance of the PDA/HA surfaces is attributable to the HA chains that probably attached onto their upper surface via hydrogen bonding between PDA and HA. At the same time, the electrostatic repulsion between PDA and HA probably prevents the aggregation of PDA, resulting in the formation of a highly uniform PDA/HA hybrid film with the HA chains (with a stretched structure) on the upper surface. We also developed a simple method for removing this PDA/HA film and recycling the Au substrates by using an aqueous solution of NaOH as the hydrolyzing agent. The Au surface remained undamaged, and a PDA/HA film could be redeposited on the surface, with the surface exhibiting good antifouling performance even after 10 such cycles. Finally, it was found that this grafting method is applicable to other substrates, including epoxy resins, polystyrene, glass, and steel, owing to the strong adhesion of PDA with these substrates.

•An electroless-plated gold film was applied to fabricate a sensitive and stable optical fiber SPR sensor.•The gold film was functionalized by the spontaneous polymerization of dopamine.•Antibody immobilization on PDA-modified gold film was very effective.•The PDA- modified gold film displayed a high sensitivity in immunoassays.A sensitive and stable electroless-plated gold film for the preparation of an optical fiber surface plasmon resonance (SPR) sensor is presented in this work, together with a facile antibody immobilization method. Gold nanoparticles were uniformly adsorbed onto the surface of an optical fiber forming a film with a thickness of approximately 56.3 nm. The sensor had a high sensitivity with 2054 nm/RIU and 3980 nm/RIU in the refractive index ranges of 1.333–1.359 and 1.359–1.386, respectively. An SPR biosensor was developed based on polydopamine-modified gold film (PDA-Au), which was fabricated by a simple and quick spontaneous polymerization of dopamine (DA) on the gold film. When goat anti-human IgG antibodies were immobilized, the PDA-Au surface had a larger resonant wavelength shift of 66.21 nm compared with the traditional 11-mercaptoundecanoic acid-modified gold film (MUA-Au) surface. In addition, the PDA-Au surface enabled the sensitive and selective determination of human IgG down to a concentration of 2 μg mL−1 with a high sensitivity of 0.41 nm per μg mL−1. The PDA-Au surface exhibited an approximately four fold higher sensitivity and an about seven fold lower LOD than the MUA-Au surface to human IgG.

A fast and facile method of fabricating fiber-optic localized surface plasmon resonance sensors based on spherical gold nanoparticles was introduced in this study. The gold nanoparticles with an average diameter of 55 nm were synthesized via the Turkevich method and were then immobilized onto the surface of an uncladded sensor probe using a polydopamine layer. To obtain a sensor probe with high sensitivity to changes in the refractive index, a set of key optimization parameters, including the sensing length, coating time of the polydopamine layer, and coating time of the gold nanoparticles, were investigated. The sensitivity of the optimized sensor probe was 522.80 nm per refractive index unit, and the probe showed distinctive wavelength shifts when the refractive index was changed from 1.328 6 to 1.398,7. When stored in deionized water at 4 °C, the sensor probe proved to be stable over a period of two weeks. The sensor also exhibited advantages, such as low cost, fast fabrication, and simple optical setup, which indicated its potential application in remote sensing and real-time detection.

Antifouling surfaces capable of reducing nonspecific protein adsorption from natural complex media are highly desirable in surface plasmon resonance (SPR) biosensors. A new protein-resistant surface made through the chemical grafting of easily available hyaluronic acid (HA) onto gold (Au) substrate demonstrates excellent antifouling performance against protein adsorption. AFM images showed the uniform HA layer with a thickness of ∼10.5 nm on the Au surface. The water contact angles of Au surfaces decreased from 103° to 12° with the covalent attachment of a carboxylated HA matrix, indicating its high hydrophilicity mainly resulted from carboxyl and amide groups in the HA chains. Using SPR spectroscopy to investigate nonspecific adsorption from single protein solutions (bovine serum albumin (BSA), lysozyme) and complex media (soybean milk, cow milk, orange juice) to an HA matrix, it was found that ultralow or low protein adsorptions of 0.6–16.1 ng/cm2 (e.g., soybean milk: 0.6 ng/cm2) were achieved on HA-Au surfaces. Moreover, anti-BSA was chosen as a model recognition molecule to characterize the immobilization capacity and the antifouling performance of anti-BSA/HA surfaces. The results showed that anti-BSA/HA sensor surfaces have a high anti-BSA loading of 780 ng/cm2, together with achieving the ultralow (<3 ng/cm2 for lysozyme and soybean milk) or low (<17 ng/cm2 for cow milk and 10% blood serum) protein adsorptions. Additionally, the sensor chips also exhibited a high sensitivity to BSA over a wide range of concentrations from 15 to 700 nM. Our results demonstrate a promising antifouling surface using extremely hydrophilic HA as matrix to resist nonspecific adsorption from complex media in SPR biosensors.Keywords: antifouling; biosensor; hyaluronic acid; nonspecific adsorption; SPR

Facile, efficient, and robust immobilization of metal nanostructures on porous bioscaffolds is an interesting topic in materials chemistry and heterogeneous catalysis. This study reports a facile in situ method for the synthesis and immobilization of small silver nanoparticles (AgNPs) at room temperature on natural eggshell membrane (ESM), which presents interwoven fibrous structure and can be used as a unique protein-based biotemplate. Procyanidin (Pro), a typical plant polyphenol extracted from grape seeds and skins, was first grafted onto ESM fibers to serve as both reductant and stabilizer during the synthesis process. As a result, the AgNPs were facilely synthesized and robustly immobilized on the ESM fibers without additional chemical reductant or physical treatments. The morphology and microstructure of the as-prepared AgNPs@Pro-ESM composites were characterized by combined microscopy and spectroscopy technologies. The results indicate that small AgNPs with mean diameter of 2.46 nm were successfully prepared on the Pro-ESM biotemplate. The composites exhibited good catalytic activity toward the reduction of 4-nitrophenol (4-NP). More importantly, these composite catalysts can be easily recovered and reused for more than eight cycles because of their high stability.Keywords: 4-nitrophenol; catalysis; eggshell membrane; procyanidin; silver nanoparticles;

Efficient immobilization of catalytic active metal nanoparticles into porous supporting materials is of important scientific interest in practice. We report on the fabrication of novel bionanocomposites, comprising a three-dimensional porous eggshell membrane (ESM) bioscaffold decorated with catalytic active metal (Pt, Pd) nanoparticles, to reduce highly toxic Cr(VI). Procyanidin (Pro), a natural plant polyphenol with abundant phenolic hydroxyls, was first covalently grafted on the ESM fiber surface to provide stable binding sites for chelating metal precursors. Highly dispersed Pt and Pd nanoparticles with small size were facilely generated and stably immobilized onto the surface of ESM followed by NaBH4 reduction. These metal nanoparticle-incorporating ESM composites were active heterogeneous catalysts for the reduction of toxic Cr(VI) to Cr(III) by employing formic acid as the reducing agent. Notably, it is easy to recover and recycle the catalysts, revealing the good stabilization of procyanidin-grafted ESM for nanoparticles.

Pomelo peel is an abundant pectin-rich biomass waste in China and has the potential to serve as a source of fuels and chemicals. This study reports a promising way to deal with pomelo peel waste and to utilize it as raw material for ethanol production via simultaneous saccharification and fermentation (SSF). An integrated strategy, incorporating hydrothermal treatment, multienzyme formulation, and fed-batch operation, was further developed to enhance the ethanol production. The results show that hydrothermal treatment (120 °C, 15 min) could significantly reduce the use of cellulase (from 7 to 3.8 FPU g–1) and pectinase (from 20 to 10 U g–1). A multienzyme complex, which consists of cellulase, pectinase, β-glucosidase, and xylanase, was also proven to be effective to improve the hydrolysis of pretreated pomelo peel, leading to higher concentrations of fermentative sugars (36 vs 14 g L–1) and galacturonic acid (23 vs 9 g L–1) than those with the use of a single enzyme. Furthermore, to increase the final ethanol concentration, fed-batch operation by adding fresh substrate was employed in the SSF process. A final solid loading of 25% (w/v), which is achieved by adding 15% fresh substrate to the SSF system at an initial solid loading of 10%, produced 36 g L–1 ethanol product in good yield (73.5%). The ethanol concentration is about 1.73-fold that at the maximum solid loading of 14% for batch operation, whereas both of them have a closed ethanol yield. The results indicate that the use of the fed-batch mode could alleviate the decrease in ethanol yield at high solid loading, which is caused by significant mass transfer limitation and increased inhibition of toxic compounds in the SSF process. The integrated strategy demonstrated in this work could open a new avenue for dealing with pectin-rich biomass wastes and utilization of the wastes to produce ethanol.

Hydrogen bonding is one of dominant forces in crystalline cellulose and lignocellulose, however, it is still a big challenge to evaluate the changes in the hydrogen bonding strength. Here, we reported a new method for measuring the changes in the hydrogen bonding strength of cellulose-based materials by terahertz time domain spectroscopy (THz-TDS). Avicel, corncob and their residual substrates after enzymatic hydrolysis were chosen as the targeted cellulose to demonstrate this method. THz adsorption in the range 0.5–2.5 THz and refractive index in the range 0.5–1.0 THz provided a direct signal corresponding to an increase in the hydrogen bonding strength for the residual samples (Avicel and corncob) after enzymatic hydrolysis. The THz results were further compared with those obtained from X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) analysis. As a quick and non-invasive technique, THz spectroscopy provides unique information about the changes in the hydrogen bonding strength of cellulose-based materials. Therefore, it is a promising way to directly evaluate the hydrogen bonding strength of cellulosic macromolecules.

Multifunctional nanoprobes with excellent reusable capability for practical detection can not only increase the resource utilization rate, but also reduce the discharge of toxic nanoparticles into the environment. In this paper, novel magnetic–fluorescent nanocomposites (Fe3O4@CdTe) have been successfully fabricated through layer-by-layer (LBL) self-assembly. The particles were used to develop a reusable fluorescence method to detect hydrogen peroxide and glucose with high sensitivity. The obtained core–shell Fe3O4@CdTe nanocomposites were characterized by transmission electron microscopy and fluorescence spectroscopy. The results indicated the successful formation of a CdTe shell on the surface of the magnetic Fe3O4 core. Its sensing performance towards H2O2 and glucose was then discussed in detail. The emission of the nanocomposites gradually reduced with the increasing analyte concentration. High sensitivity and good selectivity were observed from the composites. More importantly, these composites can be easily recovered and reused for several cycles due to their magnetism and high stability. Furthermore, we demonstrated that this fluorescent sensor can be used for glucose detection in human urine samples. As a multifunctional nanoplatform, the present nanoprobe holds genuine potential in future biosensing applications.

Pepsin is an active digestive enzyme present in the acidic environment of animal stomachs, and has been widely used to prepare bioactive peptides in the food industry. In this work, a simple fluorescence sensor for scissor-based detection of pepsin activity was developed by using lysozyme-stabilized gold nanoclusters (AuNCs@Lyz) in aqueous media. Under acidic conditions (pH 3.0), enzymatic digestion of AuNCs@Lyz with pepsin results in a significant decrease of fluorescence intensity. Notably, its acidic environment not only helps maintain the maximum fluorescence of gold nanoclusters, but also ensures the highest enzymatic activity of pepsin. In addition to offering high selectivity because of the unique proteolytic action of pepsin under acidic conditions, this facile method provides high sensitivity. With the sensing system, the linear range for pepsin detection is found to be 1 μg mL−1 to 100 μg mL−1, with a detection limit of 0.256 μg mL−1 at a signal-to-noise ratio of 3. Furthermore, the AuNCs@Lyz based fluorescent sensing system could find applications in highly sensitive and selective detection of pepsin in food and biological samples.

We report a green synthesis of novel gold nanoparticle–nanocluster composite nanostructures directly using trypsin as linking and reducing agents. Size exclusion chromatography (SEC) and transmission electron microscopy (TEM) reveals that the as-prepared gold nanocomposite (gold nanoparticles-trypsins-nanoclusters, GNPs-Trys-GNCs) is composed of GNPs (gold nanoparticles) with an average diameter of 5.5 nm and the GNCs (gold nanoclusters, about 1 nm)-embedded trypsins (Trys-GNCs), which are attached to the surface of the GNPs. The specific amino acids in the trypsin molecule, like cysteine, methionine, and tyrosine, combined with the unique spatial structures, enable trypsins to bind and reduce the AuCl4– ions, simultaneously forming GNPs and GNCs in one-pot synthesis. Similar to pure GNPs, the GNPs-Trys-GNCs nanocomposite also exhibits an intense surface plasmon resonance (SPR) absorbance at 520 nm. However, it shows an obvious different optical property in the interaction with Pb2+ ions. In the presence of Pb2+ ions, an increased intensity and a slight red-shift of the SPR peak for the GNPs-Trys-GNCs nanocomposite in the UV–vis spectra were observed, while a decreased intensity and large red-shift for pure GNPs were observed in previous studies. Moreover, we found that the SPR intensity linearly increased with Pb2+ concentration from 1.6 to 32.3 μM (R2 = 0.9731). In addition, high-level Pb2+ ions would induce the aggregation of GNPs-Trys-GNCs nanocomposite accompanied by the formation of precipitate. The unique structure and optical property of the GNPs-Trys-GNCs nanocomposite enable it to be used in heavy metal ions sensing and elimination.Keywords: Biomineralization; Gold nanoclusters; Gold nanoparticles; Nanocomposite; Nanostructure; Protein template;

For the first time, we demonstrated the fabrication of silver nanoparticles (NPs) in cross-linked protein crystal hybrid material with catalytic properties using a facile chemical reduction method. The macroscopic porous lysozyme crystals can be used as excellent templates for the incorporation of Ag nanoparticles. The resulting AgNP-in-lysozyme crystal composites exhibited a good catalytic activity toward nitrophenol reduction. Notably, these catalysts could be easily recovered and reused for at least five successive cycles with almost constant activity and conversion efficiency.

The interaction of procyanidins with proteins has aroused extensive attention due to its important relationship with the bioavailability and astringent property of polyphenols. In the present work, we have investigated the interactions of lysozyme with procyanidin dimer (B3) using various biophysical approaches, which aims to provide insights into the mechanism of protein/polyphenol aggregation. Procyanidin B3 spontaneously binds lysozyme, inducing the multilevel structural changes in lysozyme and the formation of insoluble complexes. The relationship between lysozyme aggregation and the loss of enzymatic activity was monitored using dynamic light scattering and fluorescence quenching. The influences of two carbohydrates (gum arabic and sucrose) on lysozyme/B3 aggregation were also studied. Gum arabic effectively inhibited the formation of insoluble aggregates, but was unable to restore the fluorescence and activity of lysozyme. However, sucrose concomitantly decreased the aggregate size with the recovery of fluorescence and lysozyme activity. These results proposed two probable mechanisms by which these two carbohydrates inhibit protein/polyphenol aggregation.Highlights► Interaction between procyanidin B3 and lysozyme was studied by various techniques. ► The loss of lysozyme activity was correlated with its structural change. ► Gum arabic and sucrose have great effects on protein/polyphenol association. ► We provided two probable mechanisms for the effects of carbohydrates.

Natural oligomeric procyanidin (OPC) with high biological and pharmacological activities was successfully used to synthesize OPC–insulin (OPC–INS) nanoparticles with different aggregation sizes for sustained and controlled delivery of hydrophilic insulin. The aggregation size of OPC–INS nanoparticles was regulated by OPC concentration, pH value, and incubation time. The fabrication mechanism would be that OPC and insulin self-assembled into a mixture of insulin aggregates via intermolecular interactions. In the self-assembly of insulin, OPC could serve both in the encompassing of insulin aggregates as a stabilizer and cross-linking different amounts of insulin aggregates into OPC–INS nanoparticles as interphase. OPC–INS nanoparticles not only demonstrated effective insulin drug loading but also exhibited aggregation-size-dependent and controlled insulin release performance in vitro. In the best case for OPC–INS nanoparticles, only ∼21% of insulin was released in 37 days. This study showed that the OPC–INS nanosystem is promising to serve as a long-acting insulin release formulation, and OPC has great potential as a drug carrier for nanomedicine.

The measurement of molecular parameters, such as molecular weight and degrees of polymerisation, in lignocellulosic feedstock are critical during the hydrolysis process, because following the changes in these parameters can provide insight into the mechanism of hydrolysis. This work employs SEC-MALLS to monitor the changes in the absolute values of these parameters of lignocellulosic samples during enzymatic hydrolysis. The lignocellulosic samples were directly dissolved in 8.0% LiCl/DMAc without any change. The number- and weight-average molecular weights of the cellulose fraction decreased initially and increased slightly thereafter, indicating that a “layer-by-layer” mechanism is adapted for enzymatic hydrolysis of the cellulose fraction of lignocellulose. Similar changes in the molecular weights of the hemicellulose fraction indicates that hemicellulose and cellulose are always together in the recalcitrant substrates during enzymatic hydrolysis. Finally, the reaction mechanism of cellulase on lignocellulose was defined by SEC-MALLS analysis combining HPLC and XRD data.

The properties of crystalline protein materials are closely linked to crystal shape. However, the effective strategies for the shape control of protein crystals are lacking. The conventional sitting-drop vapor-diffusion method was employed to investigate the influence of pH and temperature on the crystal nucleation behavior of hen egg white lysozyme. Moreover, the size distributions of protein crystals grown at different conditions were analyzed. Differential scanning calorimetry was employed to evaluate the thermal stability of lysozyme crystals. The results indicated that pH and temperature will affect the supersaturation and electrostatic interactions among protein molecules in the nucleation process. In particular, the crystals with different aspect ratios can be selectively nucleated, depending upon the choice of pH and temperature. Therefore, this study provided a simple method for obtaining shape-controlled lysozyme crystals and supplied some information on thermal behaviors of lysozyme crystals grown at different pH values.

The effective recycling of cellulase requires an in-depth understanding of cellulase adsorption and desorption. In the present study, we examined the adsorption behaviors and stabilities of cellulase at different pH values. Acidic pH (<4.8) was found to favor adsorption, whereas neutral and alkaline pH (especially pH 7 and 10) favored desorption. The influence of pH on cellulase activity was temperature dependent. Under mild conditions (e.g., pH 7 and 25 °C), the effect of pH on cellulase activity was reversible, and the cellulase activity can return to almost 100% by adjusting the pH value to 4.8. However, under severe conditions (e.g. pH 10 and 50 °C), irreversible inactivation may take place. We also explored the roles of pH and temperature in cellulase adsorption kinetics and isotherms. At pH 4.8, temperature had no remarkable effect on the adsorption capacity of the cellulases onto substrate. However, at pH 7 and 10, high temperatures lead to more cellulase desorption. Only at pH 4.8 does cellulase adsorption well fit (R2 > 0.96) the pseudo-first-order kinetic and Langmuir adsorption isotherm (R2 > 0.99) models.

Biofuels produced from lignocellulosic biomass can significantly reduce the energy dependency on fossil fuels and the resulting effects on environment. In this respect, cellulosic ethanol as an alternative fuel has the potential to become a viable energy source in the near future. Over the past few decades, tremendous effort has been undertaken to make cellulosic ethanol cost competitive with conventional fossil fuels. The pretreatment step is always necessary to deconstruct the recalcitrant structures and to make cellulose more accessible to enzymes. A large number of pretreatment technologies involving physical, chemical, biological, and combined approaches have been developed and tested at the pilot scale. Furthermore, various strategies and methods, including multi-enzyme complex, non-catalytic additives, enzyme recycling, high solids operation, design of novel bioreactors, and strain improvement have also been implemented to improve the efficiency of subsequent enzymatic hydrolysis and fermentation. These technologies provide significant opportunities for lower total cost, thus making large-scale production of cellulosic ethanol possible. Meanwhile, many researchers have focused on the key factors that limit cellulose hydrolysis, and analyzing the reaction mechanisms of cellulase. This review describes the most recent advances on process intensification and mechanism research of pretreatment, enzymatic hydrolysis, and fermentation during the production of cellulosic ethanol.

Enzymatic hydrolysis of corn stover was performed in an integrated membrane bioreactor (MBR) incorporating a 10 kDa flat sheet polysulfone membrane to increase cellulose conversion and to reduce enzyme dosage. Several pretreatment methods and semi-continuous MBR were examined to investigate their effect on the glucose yield and enzyme utilization efficiency. Compared with conventional batch reactor (CBR), cellulose conversion increased by 5% in a MBR because of the removal of glucose and cellobiose inhibitors. More than 15% increment in cellulose conversion was obtained using fed-MBR, and the reaction rate improved significantly. Enzyme utilization efficiency in a fed-batch MBR were 1.94-fold of CBR and 1.34-fold of fed-CBR for corn stover pretreated by soaking in aqueous ammonia and 3.31-fold of CBR and 1.32-fold of fed-CBR for corn stover pretreated by diluted sulfuric acid–sodium hydroxide.

Dibenzo [c,h]-[1,6] naphthyridin-6-one (ARC-111) and its analogues can exhibit potent topoisomerase I targeting and antitumor activities. Quantitative structure-activity relationship (QSAR) studies on the antitumor activity of 22 ARC-111 analogues to RPMI 8402 (expressed as pIC50) were performed by using electronic, spatial, thermodynamic, and topological descriptors based on different chemometric tools including stepwise multiple linear regression (stepwise MLR), partial least squares (PLS), and artificial neural network (ANN). QSAR equations were examined using a training set of 18 compounds and an external test set of four compounds, which was divided by cluster analysis. The ANN model was the most powerful, with a square of predictive correlation coefficient Rpred2 of 0.963 for the test set; the Rpred2 values for stepwise MLR and PLS were 0.780 and 0.729, respectively. The results obtained with these models indicated that the antitumor activity of ARC-111 analogues depends strongly on electronic factors (such as sum of E-state of H-bond acceptors, dipole moment as debyes, and total energy).

To enhance the conversion of the cellulose and hemicellulose, the corncob pretreated by aqueous ammonia soaking was hydrolyzed by enzyme complexes. The saturation limit for cellulase (Spezyme CP) was determined as 15 mg protein/g glucan (50 filter paper unit (FPU)/g glucan). The accessory enzymes (β-glucosidase, xylanase, and pectinase) were supplemented to hydrolyze cellobiose (cellulase-inhibiting product), hemicellulose, and pectin (the component covering the fiber surfaces), respectively. It was found that β-glucosidase (Novozyme 188) loading of 1.45 mg protein/g glucan [30 cellobiase units (CBU)/g glucan] was enough to eliminate the cellobiose inhibitor, and 2.9 mg protein/g glucan (60 CBU/g glucan) was the saturation limit. The supplementation of xylanase and pectinase can increase the conversion of cellulose and hemicellulose significantly. The yields of glucose and xylose enhanced with the increasing enzyme loading, but the increasing trend became low at high loading. Compared with xylanase, pectinase was more effective to promote the hydrolysis of cellulose and hemicellulose. The supplementation of pectinase with 0.12 mg protein/g glucan could increase the yields of glucose and xylose by 7.5% and 29.3%, respectively.

•The effect of agitation on enzymatic hydrolysis of corncob was investigated.•Cellulase was obviously deactivated under high agitation rate.•A control strategy with stepwise reduction in agitation rates was proposed.•Optimal strategy yields more glucose than most of continuous agitation.To ensure effective enzyme transfer and energy saving, the mixing control is a key issue for efficient hydrolysis of lignocellulose at high solid loading. The effect of agitation on enzymatic hydrolysis of pretreated corncob was investigated in this work. Cellulase was obviously deactivated under the agitated condition in the absence of substrate. Increasing from 0 rpm to 150 rpm, high agitation rates increased initial hydrolysis rates and final sugar concentration. However, the excessively high agitation rate at 200 rpm had a significant negative effect on hydrolysis efficiency at the solid loadings from 2% to 15%. Rapid mixing was required only to promote cellulase diffusion within the first 6 h of hydrolysis; afterwards, a smaller mixing energy was required to maintain high hydrolysis efficiency. Based on the hydrolysis rates at different agitation rates, a control strategy with stepwise reduction in agitation rates was proposed. After 72 h of hydrolysis, the glucose yield (54.92 g/L) using the optimal strategy approached that using high-rate continuous agitation (57.34 g/L) at a solid loading of 10% and was greater than those using low-rate continuous agitation. Thus, the stepwise reduction of agitation rate could be beneficial because it could save energy while producing a high sugar concentration, especially for a high solid loading reaction system.

•We designed a novel fractionation method for components of lignocellulose in corncob.•It has advantages: modest condition, low usage of alkaline, non-toxic byproduct, etc.•The recovery rates of lignin and hemicellulose were 77.5% and 89%, respectively.•We characterized the properties of three components for value-added use potential.The biorefinery process for lignocellulose conversion generally requires fractionation of its three major components including cellulose, hemicellulose, and lignin. The present work demonstrates that the alkaline hydrogen peroxide (AHP) method for fractionating lignocelluloses, such as corncob, is advantageous due to its modest reaction conditions, low alkaline usage, and effective fractionation of three components. The removal ratio of lignin and hemicellulose after 6 h treatment reached 75.4% and 38.7%, respectively, leaving the recovery ratio of cellulose in the AHP residues at 81.3%. After 24 h enzymatic hydrolysis of the AHP residues, we found that the degree of cellulose conversion was approximately 80%. Through ethanol precipitation and desalination, 89% of dissolved hemicellulose was recovered as white solids, whereas 77.5% of soluble lignin was obtained as brown solids. The isolated three components were further characterized. The major pyrolysis temperature of the AHP residues increased. And the recovered lignin revealed an increase in carboxylic acid content and a decrease in phenolic hydroxyl content by oxidation of H2O2.

•A facile method was developed for simultaneous detection of CAP and GEN using gradient elution.•The simultaneous detection was realised using SPR biosensor in a single test channel.•Alkaline solution and Gly-HCl solutions eluted GEN-Ab and CAP-Ab, respectively.•This assay exhibited high efficiency and sensitivity.For the analysis of massive samples containing multiple analytes, the enhancement of detection efficiency is crucial. In this study, a facile method was developed for sequential detection of chloramphenicol (CAP) and gentamicin (GEN) in complex samples, e.g. milk, using a surface plasmon resonance (SPR)-based biosensor. Based on the immune inhibition format, two conjugates of antigen and bovine serum albumin (BSA)—denoted as CAP–BSA and GEN–BSA—were grafted on the same channel of the SPR sensor chip. Two standard curves for CAP and GEN were separately obtained by first mixing a single antibody with different concentrations of the relevant antigen. Moreover, different regeneration solutions were screened for sequential analysis. An alkaline solution was found to completely remove the antibody against GEN (AbGEN) from the chip, but it exhibited limited ability to dissociate the antibody against CAP (AbCAP). Therefore, alkaline solution and Gly-HCl solutions are successively applied to elute AbGEN and AbCAP, respectively. By gradual elutions, CAP and GEN concentrations were simultaneously calculated with limit of detection values of 5.28 ng/mL and 2.26 ng/mL, respectively. Furthermore, the spiking milk samples with CAP and GEN validated the assay with recoveries of 77.6–101.1%. Therefore, this method is expected to improve the detection efficiency of SPR biosensors.

For the first time, we demonstrated the fabrication of silver nanoparticles (NPs) in cross-linked protein crystal hybrid material with catalytic properties using a facile chemical reduction method. The macroscopic porous lysozyme crystals can be used as excellent templates for the incorporation of Ag nanoparticles. The resulting AgNP-in-lysozyme crystal composites exhibited a good catalytic activity toward nitrophenol reduction. Notably, these catalysts could be easily recovered and reused for at least five successive cycles with almost constant activity and conversion efficiency.

In this study, we proposed a bioinspired approach for the deposition and zwitteration of hyaluronic acid (HA) with a reduced glutathione (GSH) to form a composite layer that functions as a low fouling coating. A polyanion of the HA–dopamine conjugate (HADA) possessing catechol groups was synthesized by carbodiimide chemistry between HA and dopamine. Then, the dopamine conjugated biofunctional polymers (HADA) were grafted onto Au substrates via the transformation of catechol into a quinone group under mild oxidative conditions followed by a reaction with GSH to avoid undesired adhesion and also to shield the exposed Au substrate. Analysis of XPS spectra and wettability indicated that HADA and GSH were successfully grafted onto Au substrates. Surface plasmon resonance analysis showed that both HADA and further GSH modified surfaces exhibited reduced nonspecific adsorption. The attachment of GSH to HADA modified surfaces (HADA-G) resulted in better antifouling performance, with a low or ultralow protein adsorption of 0–7.51 ng cm−2 when exposed to single protein solutions, and a reduction in nonspecific adsorption from cow's milk to 10% compared to that of bare gold. The enhanced antifouling performance of HADA-G modified surfaces was likely due to the zwitterionic structure in GSH, which can induce stronger surface hydration through electrostatic interactions as well as the hydrogen bonding induced by HADA. Our results provide a facile and universal approach to surface modification and demonstrate the benefits of using a composite layer for the design of low fouling surfaces.

This study presents a facile, rapid and effective method for the fabrication of optical fiber surface plasmon resonance (SPR) sensors via polydopamine (PDA)-accelerated electroless plating (ELP). The bioinspired PDA coating formed through the facile self-polymerization of dopamine (DA) was utilized as a versatile material for optic-fiber functionalization. Gold seeds were then rapidly and firmly adsorbed onto the PDA functional layer by amino and imino intermediates generated during the polymerization, and a gold film sensor was fabricated after metal deposition. The fabrication time of the sensor was decreased by 6–12 times, and the fabricated sensor exhibited higher sensitivity, better reproducibility and adhesion stability compared with those fabricated by the traditional ELP. Some key experimental parameters, including DA polymerization temperature, DA polymerization time, and plating time, were investigated in detail. The optimized sample exhibited high sensitivity ranging from 1391 nm per RIU to 5346 nm per RIU in the refractive index range of 1.328 to 1.386. Scanning electron microscopy images indicated that the sensor surface consisted of gold nanoparticles with a uniform particle size and an orderly arrangement, and the film thickness was approximately 60 nm. Another PDA layer was formed on the gold film for facile immobilization of antibodies. The sensor exhibited effective antibody immobilization ability and high sensitivity for human IgG detection over a wide range of concentrations from 0.5 to 40 μg mL−1, which indicate the potential applications of the fabricated sensor in immunoassays.

Natural oligomeric procyanidin (OPC) with high biological and pharmacological activities was successfully used to synthesize OPC–insulin (OPC–INS) nanoparticles with different aggregation sizes for sustained and controlled delivery of hydrophilic insulin. The aggregation size of OPC–INS nanoparticles was regulated by OPC concentration, pH value, and incubation time. The fabrication mechanism would be that OPC and insulin self-assembled into a mixture of insulin aggregates via intermolecular interactions. In the self-assembly of insulin, OPC could serve both in the encompassing of insulin aggregates as a stabilizer and cross-linking different amounts of insulin aggregates into OPC–INS nanoparticles as interphase. OPC–INS nanoparticles not only demonstrated effective insulin drug loading but also exhibited aggregation-size-dependent and controlled insulin release performance in vitro. In the best case for OPC–INS nanoparticles, only ∼21% of insulin was released in 37 days. This study showed that the OPC–INS nanosystem is promising to serve as a long-acting insulin release formulation, and OPC has great potential as a drug carrier for nanomedicine.

Pepsin is an active digestive enzyme present in the acidic environment of animal stomachs, and has been widely used to prepare bioactive peptides in the food industry. In this work, a simple fluorescence sensor for scissor-based detection of pepsin activity was developed by using lysozyme-stabilized gold nanoclusters (AuNCs@Lyz) in aqueous media. Under acidic conditions (pH 3.0), enzymatic digestion of AuNCs@Lyz with pepsin results in a significant decrease of fluorescence intensity. Notably, its acidic environment not only helps maintain the maximum fluorescence of gold nanoclusters, but also ensures the highest enzymatic activity of pepsin. In addition to offering high selectivity because of the unique proteolytic action of pepsin under acidic conditions, this facile method provides high sensitivity. With the sensing system, the linear range for pepsin detection is found to be 1 μg mL−1 to 100 μg mL−1, with a detection limit of 0.256 μg mL−1 at a signal-to-noise ratio of 3. Furthermore, the AuNCs@Lyz based fluorescent sensing system could find applications in highly sensitive and selective detection of pepsin in food and biological samples.

Multifunctional nanoprobes with excellent reusable capability for practical detection can not only increase the resource utilization rate, but also reduce the discharge of toxic nanoparticles into the environment. In this paper, novel magnetic–fluorescent nanocomposites (Fe3O4@CdTe) have been successfully fabricated through layer-by-layer (LBL) self-assembly. The particles were used to develop a reusable fluorescence method to detect hydrogen peroxide and glucose with high sensitivity. The obtained core–shell Fe3O4@CdTe nanocomposites were characterized by transmission electron microscopy and fluorescence spectroscopy. The results indicated the successful formation of a CdTe shell on the surface of the magnetic Fe3O4 core. Its sensing performance towards H2O2 and glucose was then discussed in detail. The emission of the nanocomposites gradually reduced with the increasing analyte concentration. High sensitivity and good selectivity were observed from the composites. More importantly, these composites can be easily recovered and reused for several cycles due to their magnetism and high stability. Furthermore, we demonstrated that this fluorescent sensor can be used for glucose detection in human urine samples. As a multifunctional nanoplatform, the present nanoprobe holds genuine potential in future biosensing applications.