J-Aggregate Promoting NIR-II Emission for Fluorescence/Photoacoustic Imaging-Guided Phototherapy.

J-aggregate is a promising strategy to enhance second near-infrared window (NIR-II) emission, while the controlled synthesis of J-aggregated NIR-II dyes is a huge challenge because of the lack of molecular design principle. Herein, bulk spiro[fluorene-9,9'-xanthene] functionalized benzobisthiadiazole-based NIR-II dyes (named BSFX-BBT and OSFX-BBT) are synthesized with different alkyl chains. The weak repulsion interaction between the donor and acceptor units and the S…N secondary interactions make the dyes to adopt a co-planar molecular conformation and display a peak absorption >880 nm in solution. Importantly, BSFX-BBT can form a desiring J-aggregate in the condensed state, and femtosecond transient absorption spectra reveal that the excited states of J-aggregate are the radiative states, and J-aggregate can facilitate stimulated emission. Consequently, the J-aggregated nanoparticles (NPs) display a peak emission at 1124 nm with a high relative quantum yield of 0.81%. The efficient NIR-II emission, good photothermal effect, and biocompatibility make the J-aggregated NPs demonstrate efficient antitumor efficacy via fluorescence/photoacoustic imaging-guided phototherapy. The paradigm illustrates that tuning the aggregate states of NIR-II dye via spiro-functionalized strategy is an effective approach to enhance photo-theranostic performance.

Planar-structured thiadiazoloquinoxaline-based NIR-II dye for tumor phototheranostics.

Second near-infrared (NIR-II) fluorescence imaging shows huge application prospects in clinical disease diagnosis and surgical navigation, while it is still a big challenge to exploit high performance NIR-II dyes with long-wavelength absorption and high fluorescence quantum yield. Herein, based on planar π-conjugated donor-acceptor-donor systems, three NIR-II dyes (TP-DBBT, TP-TQ1, and TP-TQ2) were synthesized with bulk steric hindrance, and the influence of acceptor engineering on absorption/emission wavelengths, fluorescence efficiency and photothermal properties was systematically investigated. Compared with TP-DBBT and TP-TQ2, the TP-TQ1 based on 6,7-diphenyl-[1,2,5]thiadiazoloquinoxaline can well balance absorption/emission wavelengths, NIR-II fluorescence brightness and photothermal effects. And the TP-TQ1 nanoparticles (NPs) possess high absorption ability at a peak absorption of 877 nm, with a high relative quantum yield of 0.69% for large steric hindrance hampering the close π-π stacking interactions. Furthermore, the TP-TQ1 NPs show a desirable photothermal conversion efficiency of 48% and good compatibility. In vivo experiments demonstrate that the TP-TQ1 NPs can serve as a versatile theranostic agent for NIR-II fluorescence/photoacoustic imaging-guided tumor phototherapy. The molecular planarization strategy provides an approach for designing efficient NIR-II fluorophores with extending absorption/emission wavelength, high fluorescence brightness, and outstanding phototheranostic performance.

Achieving cinnamic acid amides in water by a variant of acyltransferase from Mycobacterium smegmatis and its immobilized form using Ni-NTA modified aspen powder as a carrier.

An aqueous N-acylation reaction for preparing cinnamic acid amides was realized by using a variant of acyltransferase from Mycobacterium smegmatis (MsAcT-L12A), whereas the wild-type MsAcT showed no activity. MsAcT-L12A exhibited broad substrate adaptability, and preferred the substrates with electron-donating group. When the vinyl cinnamate (1a, 40 mM) and p-methoxyaniline (2a, 4 mM) were involved in the reaction, the excellent yield reached to 86.7 % ± 2.1 % within 3 h by MsAcT-L12A (1 mgpro./mL) in a PBS buffer (100 mM, pH 8.0) at 25 °C. The aqueous N-acylation reaction could be further improved by using an immobilized MsAcT-L12A. The biomass aspen powder (AP) as a carrier provided a low-cost, green, and environmental-friendly immobilization strategy. After it was modified by Ni-NTA, the obtained Ni-NAP could realize one-step purification and immobilization of MsAcT-L12A. The accomplished MsAcT-L12A-Ni-NAP exhibited excellent stability and recyclability, and retained its relative yield as 83.3 % ± 2.2 % even after the 7th cycle of reuse. Using only PBS buffer as a reaction medium, the operation for MsAcT-L12A-catalyzed acyl transfer was greatly simplified, and the improved stabilities of MsAcT-L12A-Ni-NAP could enhance its application potential.

Semi-rational engineering an aldo-keto reductase for stereocomplementary reduction of α-keto amide compounds.

Enantio-pure α-hydroxy amides are valuable intermediates for the synthesis of chiral pharmaceuticals. The asymmetric reduction of α-keto amides to generate chiral α-hydroxy amides is a difficult and challenging task in biocatalysis. In this study, iolS, an aldo-keto reductase from Bacillus subtilis 168 was exhibited as a potential biocatalyst, which could catalyze the reduction of diaryl α-keto amide such as 2-oxo-N, 2-diphenyl-acetamide (ONDPA) with moderate S-selectivity (76.1%, ee) and 60.5% conversion. Through semi-rational engineering, two stereocomplementary variants (I57F/F126L and N21A/F126A) were obtained with ee value of 97.6% (S) and 99.9% (R) toward ONDPA (1a), respectively, delivering chiral α-hydroxy amide with > 98% conversions. Moreover, the excellent S- and R-preference variants displayed improved stereoselectivities toward the other α-keto amide compounds. Molecular dynamic and docking analysis revealed that the two key residues at 21 and 126 were identified as the "switch", which specifically controlled the stereopreference of iolS by regulating the shape of substrate binding pocket as well as the substrate orientation. Our results offer an effective strategy to obtain α-hydroxy amides with high optical purity and provide structural insights into altering the stereoselectivity of AKRs.

Revisiting molecularly conformation-planarized organic dyes for NIR-II fluorescence imaging.

Fluorescence imaging in the second window (NIR-II, 1000-1700 nm) provides deeper penetration depth and higher resolution, but there is still a dilemma for designing NIR-II dyes for simultaneously enhancing fluorescence efficiency and prolonging excitation wavelength. Herein, a molecular conformation planarization strategy has been revisited to guide the synthesis of two donor-acceptor-donor dyes (named T-BBT and BT-BBT). On the one hand, conformational planarization can extend the absorption peaks of T-BBT and BT-BBT to the NIR region with high molar extinction coefficients of 30.5 × 103 and 16.4 × 103 L (mol-1 cm-1) at 1064 nm, respectively. On the other hand, structural rigidity can weaken electronic vibration coupling-related non-radiative decay pathways, whereby both T-BBT and BT-BBT display rather high fluorescence efficiencies of 3.6% and 13.5% in solution. Furthermore, a molecular doping strategy is adopted to alleviate fluorescence quenching in the aggregated state by suppressing long-distance energy migration, and 2.5 wt% doped BT-BBT nanoparticles show a high fluorescence efficiency of 2.0%, which enables the application of in vivo deep NIR-II fluorescence imaging for vessels and tumors with high resolution under 980 nm excitation. This work demonstrates that organic dyes with structural planarization can bridge the gap between NIR-II absorption and fluorescence efficiency.

2-Arachidonoylglycerol Attenuates Myocardial Fibrosis in Diabetic Mice Via the TGF-ß1/Smad Pathway.

PURPOSE: Diabetic cardiomyopathy (DM) is the cause of late cardiac dysfunction in diabetic patients. Myocardial fibrosis is the main pathological mechanism, and it is associated with transforming growth factor-ß1(TGF-ß1) expression up-regulation. 2-Arachidonoylglycerol (2-AG) is an endogenous cannabinoid that can effectively improve myocardial cell energy metabolism and cardiac function. Here, we evaluated the protective effect of 2-AG on diabetic cardiomyopathy. METHODS: Male C57BL/6 mice were injected with 2-AG intraperitoneally for 4 weeks (10 micro g/kg/day) after 12 weeks of diabetic modeling. After 4 weeks, heart function was evaluated by echocardiography. Heart structure was assessed by hematoxylin and eosin staining. Cardiac fibrosis was analyzed using immunohistochemistry, Sirius red stain, and western blot. RESULTS: After modeling in diabetic mice, cardiac ultrasonography showed decreased cardiac function and pathological findings showed myocardial fibrosis. 2-AG could effectively inhibit the up-regulation of TGF-ß1 and Smad2/3, reduce myocardial fibrosis, and ultimately improve cardiac function in diabetic mice. CONCLUSION: 2-AG reduces cardiac fibrosis via the TGF-ß1/Smad2/3 pathway and is a potential pathway for the treatment of cardiac dysfunction in diabetic mice.

Engineering silica encapsulated composite of acyltransferase from Mycobacterium smegmatis and MIL-88A: A stability-and activity-improved biocatalyst for N-acylation reactions in water.

Here the metal-organic framework material MIL-88A is used to purify and immobilize an acyltransferase from Mycobacterium smegmatis (MsAcT) simultaneously from the broken bacterial liquid. Regarding the possibility that the MsAcT@MIL-88A may display weak stability in its application, a silica layer is further introduced around it as a "shield" to protect the enzyme from degradation. The obtained MsAcT@MIL-88A@silica can exhibit high activity recovery, excellent thermal, pH, and storage stabilities compared with those of MsAcT@MIL-88A. The MsAcT@MIL-88A@silica can also be effectively recycled, and its initial activity of 84.0 % ± 1.2 % can be retained after the 5th cycle for N-acylation reaction in water. More importantly, the MsAcT@MIL-88A@silica can display much higher catalytic activity towards the reactions between ethyl or vinyl esters and aniline than those of free MsAcT and MsAcT@MIL-88A in aqueous media. This study provides a simple and inexpensive strategy to prepare MsAcT@MIL-88A@silica with high activity, stability, and excellent recyclability, and highlights its application potential as a biocatalyst.

Asymmetric Synthesis of Spiro[3,2'-morpholine-oxindoles] Derivatives via the [5 + 1] Annulation Reaction.

An efficacious method in which BINOL-type chiral imidodiphosphoric acid catalyzed the asymmetric [5 + 1] annulation reaction of 2-pyrrolylphenol with 1-methylindoline-2,3-dione was established. The strategy tolerated a broad substrate scope, and 30 examples were obtained. A range of enantioenriched spiro[3,2'-morpholine-oxindole] derivatives which incorporate a tertiary stereocenter, with moderate to excellent yields (up to 96%) and enantioselectivities (up to 99%) under mild conditions, was delivered.

Transmuscular quadratus lumborum block versus oblique subcostal transversus abdominis plane block for analgesia in laparoscopic hysterectomy: a randomised single-blind trial.

OBJECTIVE: The transmuscular quadratus lumborum (TQL) block and the oblique subcostal transversus abdominis plane (OSTAP) block both contribute to multimodal analgesia after laparoscopic surgery. The objective of this study was to compare the analgesic effects of the TQL block versus OSTAP block after laparoscopic hysterectomy. DESIGN: Prospective single-centre randomised single-blind trial. SETTING: University-affiliated hospital. PARTICIPANTS: Patients aged between 18 and 65 years scheduled for laparoscopic hysterectomy. INTERVENTIONS: Patients were randomised into two groups (1:1 ratio) and received bilateral TQL block or bilateral OSTAP block with 0.375% ropivacaine 20 mL on each side before surgery. PRIMARY AND SECONDARY OUTCOME MEASURES: The primary outcome measure was the cumulative morphine dose in the first 24 hours. The secondary outcome measures were the morphine consumption at each time interval after surgery, the time from the end of surgery to the first need for morphine, the Numerical Rating Scale (NRS) scores for visceral and incisional pain intensity, and the incidence of adverse events. RESULTS: The cumulative morphine dose was significantly lower in the TQL group than in the OSTAP group (17.2 (12.5) vs 26.1 (13.3) mg, p=0.010). Compared with the OSTAP group, the morphine doses from 6 to 12, 12 to 18, and 18 to 24 hours were significantly lower, the time of first need for morphine was significantly longer and the NRS scores for visceral pain intensity were significantly lower in the TQL group. CONCLUSION: Compared with the OSTAP block, the TQL block reduced morphine consumption and provided better visceral pain relief with a longer duration of effect after laparoscopic hysterectomy. TRIAL REGISTRATION NUMBER: Chinese Clinical Trial Registry (ChiCTR1800017995); pre-results.

Comparison of the effect of different volumes ropivacaine on deep serratus anterior plane block in patients undergoing breast surgery: a prospective randomized double-blinded trial.

BACKGROUND: A larger volume of local anesthetic provides a wider range of blocked sensory but carries a greater risk. The purpose of this trial was to compare the effect of different volumes of ropivacaine injected to deep serratus anterior plane in patients undergoing breast surgery. METHODS: In this randomized double-blind trial, 60 patients undergoing breast surgery were randomly allocated to R10, R20 and R30 groups (n=20), and received deep serratus anterior plane block with 10, 20 and 30 mL of 0.5% ropivacaine respectively. 30 minutes after block, the cutaneous sensory was tested by cold stimulus in the craniocaudal direction along the midaxillary line. We recorded the numerical rating scale pain scores over 24 h after surgery and estimated the area under curve by numerical rating scale pain scores. The cases of rescue analgesia and the prevalence of adverse events were also recorded. RESULTS: The blocked dermatomes were 3 [3, 4], 6 [5, 7] and 7 [6, 8] in the R10, R20 and R30 groups, respectively (R10 vs. R20, P<0.001; R10 vs. R30, P<0.001; R20 vs. R30, P=0.005). The area under curve of R10 group was significantly higher compared with the R20 and R30 groups (P=0.014, P=0.003, at rest; P<0.001, P<0.001, on movement). CONCLUSIONS: The blocked dermatomes increased with increasing volume when 10, 20 and 30 mL ropivacaine was used for deep serratus anterior plane block. The analgesic effects of 20 and 30 mL were similar to each other and better than 10 mL. Therefore, in breast surgery, volume of 20 mL ropivacaine is considered to be appropriate for deep serratus anterior plane block.

Comparison of Effectiveness of Ropivacaine Infusion Regimens for Continuous Femoral Nerve Block for Recovery After Total Knee Arthroplasty: A Randomized Double-Blind Trial.

PURPOSE: Continuous femoral nerve block (cFNB) is effective for analgesia after total knee arthroplasty (TKA). However, it is not clear which low-dose regimen of ropivacaine infusion for cFNB provides adequate analgesia and enables rapid recovery. The aim of this study was to compare the effects of different cFNB regimens on rehabilitation of patients after TKA. PATIENTS AND METHODS: Sixty patients scheduled for TKA were enrolled in this trial. After surgery, patients in the 0.1%, 0.15%, and 0.2% groups received infusion of 10 mL of 0.1%, 6.7 mL of 0.15%, and 5 mL of 0.2% ropivacaine per hour, respectively (n=20), at the dose of 10 mg/h for 48 h. The primary endpoint was time to readiness for discharge. The secondary endpoints were time to first walk, manual muscle testing (MMT) scores, numerical rating scale (NRS) scores at rest and movement, morphine consumption, rescue analgesia, and the incidence of adverse events. RESULTS: The time to readiness for discharge and the time to first walk of the 0.1% group were significantly longer than that of the 0.15% and 0.2% groups. MMT scores of the 0.2% group at 18 h after surgery were significantly lower than those of the 0.1% group. MMT scores of the 0.2% group at 24 and 48 h after surgery were also significantly lower than those of the 0.1% and 0.15% groups. NRS scores at rest and at movement in the 0.1% group were significantly higher than those in the 0.15% and 0.2% groups. CONCLUSION: Patients administered the regimens of 0.15% and 0.2% ropivacaine infusion for cFNB were ready for discharge earlier than the 0.1% group after TKA, at the dose of 10 mg/h for 48 h. The regimen of 0.15% ropivacaine, which is associated with less quadriceps muscle strength weakness than 0.2%, is recommended for postoperative analgesia after TKA.

Effects of Ropivacaine Concentration on Analgesia After Ultrasound-Guided Serratus Anterior Plane Block: A Randomized Double-Blind Trial.

PURPOSE: Serratus anterior plane (SAP) block is effective for analgesia after breast surgery. Whether a higher local anesthetic concentration prolongs sensory block duration and improves postoperative analgesia remains unclear. The aim of this study was to compare the analgesic effects of SAP block with different concentrations of ropivacaine. PATIENTS AND METHODS: Sixty patients scheduled for breast surgery were enrolled in this randomized double-blind trial. SAP block was induced with 20 mL of 0.375%, 0.5%, or 0.75% ropivacaine in Group R0.375, Group R0.5, and Group R0.75, respectively. The primary endpoint was the area under the curve (AUC) of numerical rating scale (NRS) pain intensity scores at rest over time. The secondary endpoints were AUC of NRS pain intensity scores on movement over time, active sensory block duration, tramadol consumption, and the elapsed time between completion of surgery and the first administration of rescue analgesia. RESULTS: The AUC of NRS pain intensity scores at rest of Group R0.375 was significantly higher than that of Groups R0.5 and R0.75 (P=0.025, and P=0.001). The AUC of NRS pain intensity scores on movement of Group R0.375 was also significantly higher than that of Groups R0.5 and R0.75 (both P<0.001). At higher ropivacaine concentrations, the duration of SAP sensory block increased (P<0.001). Tramadol consumption and the elapsed time between completion of surgery and the first administration of rescue analgesia were similar in the three groups (P>0.05). CONCLUSION: A comparison of 0.5% and 0.75% ropivacaine showed no significant difference in postoperative analgesia, but both were superior to 0.375% ropivacaine, although higher ropivacaine concentration lengthened the duration of SAP block. Therefore, SAP block with 0.5% ropivacaine is recommended for postoperative analgesia in breast surgery.

Rational enhancement of enzyme-catalyzed enantioselective reaction by construction of recombinant enzymes based on additive strategy.

A rational enhancement of kinetic resolution process for producing (S)-N-(2-ethyl-6-methylphenyl) alanine from racemic methyl ester using lipase B from Candida antarctica (CalB) was investigated. With the benefit results that lipase CalB-catalyzed reactions can be effectively regulated using amino acids (such as histidine and lysine) as additives, CalBs modified (mCalBs) by n-histidines at the N terminal and n-lysines at the C terminal were constructed and expressed. The results show that both soluble and precipitated mCalBs can effectively catalyze the hydrolysis reaction without adding any extra additives. The enantioselective ratio (E value) of soluble and precipitated mCalBs could be improved from 12.1 to 20.3, which were higher than that (E value was only 10.2) of commercial Novozym 435 (immobilized CalB). The study indicated that the amino acid-rich molecules introduced on lipase CalB can produce positive effects on enantioselectivity of enzyme. It provides unusual ideas for reasonable regulation of enzyme-catalyzed reactions.

Directly covalent immobilization of Candida antarctica lipase B on oxidized aspen powder by introducing poly­lysines: An economical approach to improve enzyme performance.

In our previous study, we could achieve high soluble expression of Candida antarctica lipase B (CalB) in E. coli by fusion poly­amino acid tags on CalB (pCalB). Herein, we are surprised to find that pCalB can be easily and directly covalent binding on a simply oxidized aspen powder (OAP) by the aid of poly­lysine tags. Under the optimal conditions, 72.9 ±â€¯3.6% of the total protein could be immobilized, and the activity recovery of immobilized pCalB (pCalB-OAP) was 98.9 ±â€¯3.8%. The analysis of scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR) indicated that OAP was a suitable carrier for enzyme immobilization. The immobilized pCalB-OAP could exhibit excellent thermal stabilities, and it retained a residual activity of 58.4 ±â€¯2.8% at 55 °C, whereas only 21.2 ±â€¯2.2% of its initial activity for free pCalB was observed. And it could also display a nice tolerance for the changes of pH environment, compared with that of free pCalB. The results that pCalB-OAP could retained 73.6 ±â€¯2.9% of their initial activity in (R, S)-NEMPAME hydrolysis after the tenth cycles, suggested that pCalB-OAP could be effectively recycled. The immobilization strategies established here were simple and inexpensive.

Functionalized Poplar Powder as a Support Material for Immobilization of Enoate Reductase and a Cofactor Regeneration System.

In this study, functionalized poplar powder (FPP) was used as a support material for the immobilization of enoate reductase (ER) and glucose-6-phosphate dehydrogenase (GDH) by covalent binding. Under optimal conditions, the immobilization efficiency of ER-FPP and GDH-FPP was 95.1% and 84.7%, and the activity recovery of ER and GDH was 47.5% and 37.8%, respectively. Scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS) analysis indicated that FPP was a suitable carrier for enzyme immobilization. ER-FPP and GDH-FPP exhibit excellent thermal stabilities and superior reusability. Especially, ER-FPP and GDH-FPP enable the continuous conversion of 4-(4-Methoxyphenyl)-3-buten-2-one with NAD+ recycling. While the immobilization strategies established here were simple and inexpensive, they exploited a new method for the immobilization and application of ER and its cofactor recycling system.

Stereoselective synthesis of a key chiral intermediate of (S)-Rivastigmine by AKR-GDH recombinant whole cells.

Herein, three aldo/ketoreductases (AKRs) were obtained and used to prepare N- ethyl-methyl-carbamic acid-3-[(1S)-hydroxy-ethyl]-phenyl ester ((S)-NEMCA-HEPE), which is a key intermediate of (S)-Rivastigmine. To avoid the usage of extra cofactors, the recombinant whole-cells containing AKRs and glucose dehydrogenase (GDH) were constructed and applied in the reduction reaction. The excellent conversion of 83.3% and enantiomeric excess (e.e.p) of 99.9% for (S)-NEMCA-HEPE was obtained when the recombinant whole cell (iolS-GDH) was selected as a catalyst. Additional introduction of 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIm]BF4) in the whole cell-catalyzed reaction system, the reaction rates can be further enhanced, and the reaction conversion can be increased to 98.3% only in 1 h. The analysis of flow cytometry (FCM) and ultraviolet spectrum shows that [BMIm]BF4 can greatly enhance the whole cell-catalyzed reaction activities by affecting the permeability of cell membrane. This study provided a low-cost and efficient process for preparation of (S)-NEMCA-HEPE with high optical purity.

Co-immobilization of enoate reductase with a cofactor-recycling partner enzyme.

Herein we established co-immobilized methods for enoate reductases (ERs) and glucose dehydrogenase (GDH), forming a cofactor regeneration system. In cross-linked enzyme aggregates (CLEAs), ammonium sulfate and oxidized dextran were selected as a precipitant and a cross-linker, respectively. In biomimetic immobilization (BI), ER-GDH-silica particles (ER-GDH-SPs) were rapidly formed through a one-step approach by using a silicic acid precursor. Under the optimal conditions, the ER activity recovery in ER-GDH-CLEAs and ER-GDH-SPs were 44.9±1.8% and 44.5±2.1%, and the immobilization efficiency was 93.5±1.2% and 92.4±1.2%, respectively. ER-GDH-CLEAs and ER-GDH-SPs exhibit excellent thermal and pH stability, and superior reusability. The activity of ER-GDH-SPs toward the substrate is also better than that of free ER and GDH in reduction of 4-(4-Methoxyphenyl)-3-buten-2-one. This study introduces simple and inexpensive co-immobilization strategies to construct novel and efficient ER-GDH-CLEAs and ER-GDH-SPs with high activity and stability.

Simultaneously achieve soluble expression and biomimetic immobilization of Candida antarctica lipase B by introducing polyamine tags.

Polyamine tags fused in Candida antarctica lipase B (CalB) can help achieve high soluble expression of CalB in E. coli and can directly mediate silicification, which leads to rapid formation of a CalB-silica particle complex through a one-step approach. After optimization experiments, the fused lipase CalB tagged with 6-histidine at the N terminal and 10-lysine at the C terminal (6His-CalB-10Lys) is effectively expressed with high solubility (0.1mg/mL) and specific activity (10.1U/mg), and easily cross-linked in silica particles with a high immobilization efficiency of 96.8% and activity recovery of 81.5%. The immobilized lipase 6His-CalB-10Lys exhibits excellent performance at broad temperature ranges, high thermal and storage stabilities, and superior reusability. Michaelis-Menten kinetics indicates that the affinity and enantioselectivity of the free and immobilized 6His-CalB-10Lys toward the substrate are better than that of commercial Novozym 435 in enantioselective resolution of (S)-N-(2-ethyl-6-methylphenyl) alanine ((S)-NEMPA). The strategies described in this paper are useful for the facile expression and construction of diverse enzyme systems with high efficiency and excellent recyclability.

First Novozym 435 lipase-catalyzed Morita-Baylis-Hillman reaction in the presence of amides.

The first Novozym 435 lipase-catalyzed Morita-Baylis-Hillman (MBH) reaction with amides as co-catalyst was realized. Results showed that neither Novozym 435 nor amide can independently catalyze the reaction. This co-catalytic system that used a catalytic amount of Novozym 435 with a corresponding amount of amide was established and optimized. The MBH reaction strongly depended on the structure of aldehyde substrate, amide co-catalyst, and reaction additives. The optimized reaction yield (43.4%) was achieved in the Novozym 435-catalyzed MBH reaction of 2, 4-dinitrobenzaldehyde and cyclohexenone with isonicotinamide as co-catalyst and ß-cyclodextrin as additive only in 2 days. Although enantioselectivity of Novozym 435 was not found, the results were still significant because an MBH reaction using lipase as biocatalyst was realized for the first time.

Enzyme-Catalyzed Henry Reaction in Choline Chloride-Based Deep Eutectic Solvents.

The enzyme-catalyzed Henry reaction was realized using deep eutectic solvents (DESs) as a reaction medium. The lipase from Aspergillus niger (lipase AS) showed excellent catalytic activity toward the substrates aromatic aldehydes and nitromethane in choline chloride:glycerol at a molar ratio of 1:2. Addition of 30 vol% water to DES further improved the lipase activity and inhibited DES-catalyzed transformation. A final yield of 92.2% for the lipase AS-catalyzed Henry reaction was achieved under optimized reaction conditions in only 4 h. In addition, the lipase AS activity was improved by approximately 3-fold in a DES-water mixture compared with that in pure water, which produced a final yield of only 33.4%. Structural studies with fluorescence spectroscopy showed that the established strong hydrogen bonds between DES and water may be the main driving force that affects the spatial conformation of the enzyme, leading to a change in lipase activity. The methodology was also extended to the aza-Henry reaction, which easily occurred in contrast to that in pure water. The enantioselectivity of both Henry and aza-Henry reactions was not found. However, the results are still remarkable, as we report the first use of DES as a reaction medium in a lipase-catalyzed Henry reaction.