Enzymatic catalysis is considered to be among the most environmental friendly processes for the synthesis of fine chemicals. In this study, lipase from Thermomyces lanuginosus (Lecitase Ultra™) was used to catalyze the synthesis of flavor esters, i.e., methyl butanoate and methyl benzoate by esterification of the acids with methanol in a microfluidic system. Maximum reaction rates of 195 and 115 mM min(-1) corresponding to catalytic efficiencies (k cat/K M) of 0.30 and 0.24 min(-1) mM(-1) as well as yield conversion of 54 and 41 % were observed in methyl butanoate and methyl benzoate synthesis, respectively. Catalytic turnover (k cat) was higher for methyl butanoate synthesis. Rate of synthesis and yield decreased with increasing flow rates. For both esters, increase in microfluidic flow rate resulted in increased advective transport over molecular diffusion and reaction rate, thus lower conversion. In microfluidic synthesis using T. lanuginosus lipase, the following reaction conditions were 40 °C, flow rate 0.1 mL min(-1), and 123 U g(-1) enzyme loading found to be the optimum operating limits. The work demonstrated the application of enzyme(s) in a microreactor system for the synthesis of industrially important esters.
Growth associated biosynthesis of medium chain length poly-3-hydroxyalkanoates (mcl-PHA) in Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. Models with substrate inhibition terms described well the kinetics of its growth. Selected fatty acids (C8:0 to C18:1) and ammonium were used as carbon and nitrogen sources during growth and PHA biosynthesis, resulting in PHA accumulation of about 50 to 69% (w/w) and PHA yields ranging from 10.12 g L(-1) to 15.45 g L(-1), respectively. The monomer composition of the PHA ranges from C4 to C14, and was strongly influenced by the type of carbon substrate fed. Interestingly, an odd carbon chain length (C7) monomer was also detected when C18:1 was fed. Polymer showed melting temperature (T m) of 42.0 (± 0.2) °C, glass transition temperature (T g) of -1.0 (± 0.2) °C and endothermic melting enthalpy of fusion (ΔHf) of 110.3 (± 0.1) J g(-1). The molecular weight (M w) range of the polymer was relatively narrow between 55 to 77 kDa.
The biosynthesis and characterization of medium chain length poly-3-hydroxyalkanoates (mcl-PHA) produced by Pseudomonas putida Bet001 isolated from palm oil mill effluent was studied. The biosynthesis of mcl-PHA in this newly isolated microorganism follows a growth-associated trend. Mcl-PHA accumulation ranging from 49.7 to 68.9% on cell dry weight (CDW) basis were observed when fatty acids ranging from octanoic acid (C(8:0)) to oleic acid (C(18:1)) were used as sole carbon and energy source. Molecular weight of the polymer was found to be ranging from 55.7 to 77.7 kDa. Depending on the type of fatty acid used, the (1)H NMR and GCMSMS analyses of the chiral polymer showed a composition of even and odd carbon atom chain with monomer length of C4 to C14 with C8 and C10 as the principal monomers. No unsaturated monomer was detected. Thermo-chemical analyses showed the accumulated PHA to be semi-crystalline polymer with good thermal stability, having a thermal degradation temperature (T(d)) of 264.6 to 318.8 (± 0.2) (o)C, melting temperature (T(m)) of 43. (± 0.2) (o)C, glass transition temperature (T(g)) of -1.0 (± 0.2) (o)C and apparent melting enthalpy of fusion (ΔH(f)) of 100.9 (± 0.1) J g(-1).
Lipase-catalyzed synthesis of 6-O-glucosyldecanoate from d-glucose and decanoic acid was performed in dimethyl sulfoxide (DMSO), a mixture of DMSO and tert-butanol and tert-butanol alone with a decreasing order of polarity. The highest conversion yield (> 65%) of decanoic acid was obtained in the blended solvent of intermediate polarity mainly because it could dissolve relatively large amounts of both the reactants. The reaction obeyed Michaelis-Menten type of kinetics. The affinity of the enzyme towards the limiting substrate (decanoic acid) was not affected by the polarity of the solvent, but increased significantly with temperature. The esterification reaction was endothermic with activation energy in the range of 60-67 kJ mol⁻¹. Based on the Gibbs energy values, in the solvent blend of DMSO and tert-butanol the position of the equilibrium was shifted more towards the products compared to the position in pure solvents. Monoester of glucose was the main product of the reaction.
Ultrasonic irradiation greatly improved the Candida antarctica lipase B mediated ring opening polymerization of ε-caprolactone to poly-6-hydroxyhexanoate in the ionic liquid 1-ethyl-3-methylimidazolium tetraflouroborate. Compared to the conventional nonsonicated reaction, sonication improved the monomer conversion by 63% and afforded a polymer product of a narrower molecular weight distribution and a higher degree of crystallinity. Under sonication, the polydispersity index of the product was ~1.44 compared to a value of ~2.55 for the product of the conventional reaction. With sonication, nearly 75% of the monomer was converted to product, but the conversion was only ~16% for the reaction carried out conventionally. Compared to conventional operation, sonication enhanced the rate of polymer propagation by >2-fold and the turnover number of the lipase by >3-fold.
Four different lipases were compared for ultrasound-mediated synthesis of the biodegradable copolymer poly-4-hydroxybutyrate-co-6-hydroxyhexanoate. The copolymerization was carried out in chloroform. Of the enzymes tested, Novozym 435 exhibited the highest copolymerization rate, in fact the reaction rate was observed to increase with about 26-fold from 30 to 50°C (7.9×10(-3)Ms(-1)), sonic power intensity of 2.6×10(3)Wm(-2) and dissipated energy of 130.4Jml(-1). Copolymerization rates with the Candida antarctica lipase A, Candida rugosa lipase, and Lecitase Ultra™ were lower at 2.4×10(-4), 1.3×10(-4) and 3.5×10(-4)Ms(-1), respectively. The catalytic efficiency depended on the enzyme. The efficiency ranged from 4.15×10(-3)s(-1)M(-1) for Novozym 435-1.48×10(-3)s(-1)M(-1) for C. rugosa lipase. Depending on the enzyme and sonication intensity, the monomer conversion ranged from 8.2% to 48.5%. The sonication power, time and temperature were found to affect the rate of copolymerization. Increasing sonication power intensity from 1.9×10(3) to 4.5×10(3)Wm(-2) resulted in an increased in acoustic pressure (P(a)) from 3.7×10(8) to 5.7×10(8)Nm(-2) almost 2.4-3.7 times greater than the acoustic pressure (1.5×10(8)Nm(-2)) that is required to cause cavitation in water. A corresponding acoustic particle acceleration (a) of 9.6×10(3)-1.5×10(4)ms(-2) was calculated i.e. approximately 984-1500 times greater than under the action of gravity.
The effects of organic solvents and their binary mixture in the glucose functionalization of bacterial poly-3-hydroxyalkanoates catalyzed by Lecitase™ Ultra were studied. Equal volume binary mixture of DMSO and chloroform with moderate polarity was more effective for the enzyme catalyzed synthesis of the carbohydrate polymer at ≈38.2 (±0.8)% reactant conversion as compared to the mono-phasic and other binary solvents studied. The apparent reaction rate constant as a function of medium water activity (aw) was observed to increase with increasing solvent polarity, with optimum aw of 0.2, 0.4 and 0.7 (±0.1) observed in hydrophilic DMSO, binary mixture DMSO:isooctane and hydrophobic isooctane, respectively. Molecular sieve loading between 13 to 15gL(-1) (±0.2) and reaction temperature between 40 to 50°C were found optimal. Functionalized PHA polymer showed potential characteristics and biodegradability.
Insufficient power generation from a microbial fuel cell (MFC) hampers its progress towards utility-scale development. Electrode modification with biopolymeric materials could potentially address this issue. In this study, medium-chain-length poly-3-hydroxyalkanoates (PHA)/carbon nanotubes (C) composite (CPHA) was successfully applied to modify the surface of carbon cloth (CC) anode in MFC. Characterization of the functional groups on the anodic surface and its morphology was carried out. The CC-CPHA composite anode recorded maximum power density of 254 mW/m2, which was 15-53% higher than the MFC operated with CC-C (214 mW/m2) and pristine CC (119 mW/m2) as the anode in a double-chambered MFC operated with Escherichia coli as the biocatalyst. Electrochemical impedance spectroscopy and cyclic voltammetry showed that power enhancement was attributed to better electron transfer capability by the bacteria for the MFC setup with CC-CPHA anode.
Wound management and healing in several physiological or pathological conditions, particularly when comorbidities are involved, usually proves to be difficult. This presents complications leading to socio-economic and public health burdens. The accelerative wound healing potential of biocompatible poly(3-hydroxyalkanoates)-co-(6-hydroxyhexanoate) (PHA-PCL) composite hydrogel is reported herein. The biosynthesized PHA-PCL macromer was cross-linked with PEGMA to give a hydrogel. Twenty-four rats weighing 200-250 g each were randomly assigned to four groups of six rats. Rats in group I (negative control) were dressed with sterilized gum acacia paste in 10% normal saline while PEGMA-alone hydrogel (PH) was used to dress group II (secondary control) rats. Group III rats were dressed with PHAs-PCL cross-linked PEGMA hydrogel (PPH). For the positive control (group IV), the rats were dressed with Intrasite(®) gel. Biochemical, histomorphometric and immunohistomorphometric analyses revealed a significant difference in area closure and re-epithelialization on days 7 and 14 in PPH or Intrasite(®) gel groups compared to gum acacia or PEGMA-alone groups. Furthermore, wounds dressed with PPH or Intrasite(®) gel showed evident collagen deposition, enhanced fibrosis and extensively organized angiogenesis on day 14 compared to the negative control group. While improvement in wound healing of the PH dressed group could be observed, there was no significant difference between the negative control group and the PH dressed group in any of the tests. The findings suggested that topical application of PPH accelerated the rats' wound healing process by improving angiogenesis attributed to the increased microvessel density (MVD) and expressions of VEGF-A in tissue samples. Thus, PPH has been demonstrated to be effective in the treatment of cutaneous wounds in rats, and could be a potential novel agent in the management and acceleration of wound healing in humans and animals.
Natural biopolymers have many attractive medical applications; however, complications due to fibrosis caused a reduction in diffusion and dispersal of nutrients and waste products. Consequently, severe immunocompatibility problems and poor mechanical and degradation properties in synthetic polymers ensue. Hence, the present study investigates a novel hydrogel material synthesized from caprolactone, ethylene glycol, ethylenediamine, polyethylene glycol, ammonium persulfate, and tetramethylethylenediamine via chemo-enzymatic route. Spectroscopic analyses indicated the formation of polyurea and polyhydroxyurethane as the primary building block of the hydrogel starting material. Biocompatibility studies showed positive observation in biosafety test using direct contact cytotoxicity assay in addition to active cellular growth on the hydrogel scaffold based on fluorescence observation. The synthesized hydrogel also exhibited (self)fluorescence properties under specific wavelength excitation. Hence, synthesized hydrogel could be a potential candidate for medical imaging as well as tissue engineering applications as a tissue expander, coating material, biosensor, and drug delivery system.
Bacterial cellulose (BC) has gained attention among researchers in materials science and bio-medicine due to its fascinating properties. However, BC's fibre collapse phenomenon (i.e., its inability to reabsorb water after dehydration) is one of the drawbacks that limit its potential. To overcome this, a catalyst-free thermal crosslinking reaction was employed to modify BC using citric acid (CA) without compromising its biocompatibility. FTIR, XRD, SEM/EDX, TGA, and tensile analysis were carried out to evaluate the properties of the modified BC (MBC). The results confirm the fibre crosslinking phenomenon and the improvement of some properties that could be advantageous for various applications. The modified nanofibre displayed an improved crystallinity and thermal stability with increased water absorption/swelling and tensile modulus. The MBC reported here can be used for wound dressings and tissue scaffolding.