Autonomous closed-loop mechanistic investigation of molecular electrochemistry via automation.

Electrochemical research often requires stringent combinations of experimental parameters that are demanding to manually locate. Recent advances in automated instrumentation and machine-learning algorithms unlock the possibility for accelerated studies of electrochemical fundamentals via high-throughput, online decision-making. Here we report an autonomous electrochemical platform that implements an adaptive, closed-loop workflow for mechanistic investigation of molecular electrochemistry. As a proof-of-concept, this platform autonomously identifies and investigates an EC mechanism, an interfacial electron transfer (E step) followed by a solution reaction (C step), for cobalt tetraphenylporphyrin exposed to a library of organohalide electrophiles. The generally applicable workflow accurately discerns the EC mechanism's presence amid negative controls and outliers, adaptively designs desired experimental conditions, and quantitatively extracts kinetic information of the C step spanning over 7 orders of magnitude, from which mechanistic insights into oxidative addition pathways are gained. This work opens opportunities for autonomous mechanistic discoveries in self-driving electrochemistry laboratories without manual intervention.

Interrogating the Mechanistic Features of Ni(I)-Mediated Aryl Iodide Oxidative Addition Using Electroanalytical and Statistical Modeling Techniques.

While the oxidative addition of Ni(I) to aryl iodides has been commonly proposed in catalytic methods, an in-depth mechanistic understanding of this fundamental process is still lacking. Herein, we describe a detailed mechanistic study of the oxidative addition process using electroanalytical and statistical modeling techniques. Electroanalytical techniques allowed rapid measurement of the oxidative addition rates for a diverse set of aryl iodide substrates and four classes of catalytically relevant complexes (Ni(MeBPy), Ni(MePhen), Ni(Terpy), and Ni(BPP)). With >200 experimental rate measurements, we were able to identify essential electronic and steric factors impacting the rate of oxidative addition through multivariate linear regression models. This has led to a classification of oxidative addition mechanisms, either through a three-center concerted or halogen-atom abstraction pathway based on the ligand type. A global heat map of predicted oxidative addition rates was created and shown applicable to a better understanding of the reaction outcome in a case study of a Ni-catalyzed coupling reaction.

Three-Stage Conversion of Chemically Inert n-Heptane to α-Hydrazino Aldehyde Based on Bioelectrocatalytic C-H Bond Oxyfunctionalization.

Simple petrochemical feedstocks are often the starting material for the synthesis of complex commodity and fine and specialty chemicals. Designing synthetic pathways for these complex and specific molecular structures with sufficient chemo-, regio-, enantio-, and diastereo-selectivity can expand the existing petrochemicals landscape. The two overarching challenges in designing such pathways are selective activation of chemically inert C-H bonds in hydrocarbons and systematic functionalization to synthesize complex structures. Multienzyme cascades are becoming a growing means of overcoming the first challenge. However, extending multienzyme cascade designs is restricted by the arsenal of enzymes currently at our disposal and the compatibility between specific enzymes. Here, we couple a bioelectrocatalytic multienzyme cascade to organocatalysis, which are two distinctly different classes of catalysis, in a single system to address both challenges. Based on the development and utilization of an anthraquinone (AQ)-based redox polymer, the bioelectrocatalytic step achieves regioselective terminal C-H bond oxyfunctionalization of chemically inert n-heptane. A second biocatalytic step selectively oxidizes the resulting 1-heptanol to heptanal. The succeeding inherently simple and durable l-proline-based organocatalysis step is a complementary partner to the multienzyme steps to further functionalize heptanal to the corresponding α-hydrazino aldehyde. The "three-stage" streamlined design exerts much control over the chemical conversion, which renders the collective system a versatile and adaptable model for a broader substrate scope and more complex C-H functionalization.

Investigating Oxidative Addition Mechanisms of Allylic Electrophiles with Low-Valent Ni/Co Catalysts Using Electroanalytical and Data Science Techniques.

The catalysis by a π-allyl-Co/Ni complex has drawn significant attention recently due to its distinct reactivity in reductive Co/Ni-catalyzed allylation reactions. Despite significant success in reaction development, the critical oxidative addition mechanism to form the π-allyl-Co/Ni complex remains unclear. Herein, we present a study to investigate this process with four catalysis-relevant complexes: Co(MeBPy)Br2, Co(MePhen)Br2, Ni(MeBPy)Br2, and Ni(MePhen)Br2. Enabled by an electroanalytical platform, Co(I)/Ni(I) species were found responsible for the oxidative addition of allyl acetate. Kinetic features of different substrates were characterized through linear free-energy relationship (Hammett-type) studies, statistical modeling, and a DFT computational study. In this process, a coordination-ionization-type transition state was proposed, sharing a similar feature with Pd(0)-mediated oxidative addition in Tsuji-Trost reactions. Computational and ligand structural analysis studies support this mechanism, which should provide key information for next-generation catalyst development.

Cobalt-electrocatalytic HAT for functionalization of unsaturated C-C bonds.

The study and application of transition metal hydrides (TMHs) has been an active area of chemical research since the early 1960s1, for energy storage, through the reduction of protons to generate hydrogen2,3, and for organic synthesis, for the functionalization of unsaturated C-C, C-O and C-N bonds4,5. In the former instance, electrochemical means for driving such reactivity has been common place since the 1950s6 but the use of stoichiometric exogenous organic- and metal-based reductants to harness the power of TMHs in synthetic chemistry remains the norm. In particular, cobalt-based TMHs have found widespread use for the derivatization of olefins and alkynes in complex molecule construction, often by a net hydrogen atom transfer (HAT)7. Here we show how an electrocatalytic approach inspired by decades of energy storage research can be made use of in the context of modern organic synthesis. This strategy not only offers benefits in terms of sustainability and efficiency but also enables enhanced chemoselectivity and distinct, tunable reactivity. Ten different reaction manifolds across dozens of substrates are exemplified, along with detailed mechanistic insights into this scalable electrochemical entry into Co-H generation that takes place through a low-valent intermediate.

The efficacy of ankle arthroplasty on motor function recovery in patients with orthopedic ankle injury: a systematic review and meta-analysis.

BACKGROUND: This study analyzed the effects of ankle arthroplasty on the recovery of motor function in patients with orthopedic ankle injury. METHODS: English databases including PubMed, Web of Science, Embase, and the Cochrane Library were searched for randomized controlled trials (RCTs) examining the effects of ankle arthroplasty, ankle replacement, and joint prosthesis on motor function recovery in patients with orthopedic ankle injury. The outcome indicators included the American Orthopedic Foot and Ankle Society (AOFAS) score, the 36-item short form survey (SF-36) score, the Foot and Ankle Ability Measures (FAAM) score, and the visual analog scale (VAS) score. The quality of the included literature was evaluated using the Jadad tool, and meta-analysis of the experimental data was performed using the Review Manager 5.3 software. RESULTS: A total of 7 articles, including 443 patients, were analyzed. The meta-analysis showed significant improvement in AOFAS scores among patients in the experiment group (who underwent ankle replacement) compared with those in the control group (who did not undergo ankle replacement) [mean difference (MD) =-41.89, 95% confidence interval (CI): -51.29 to 32.49, Z=8.73, P<0.00001], VAS scores (MD =5.59, 95% CI: 4.84 to 6.34, Z=14.56, P<0.00001), SF-36 scores (MD =-13.89, 95% CI: -26.74 to 1.04, Z=2.12, P=0.03), and FAAM scores (MD =-25.78, 95% CI: -31.27 to 20.29, Z=9.20, P<0.00001) compared to patients in the control group. DISCUSSION: Ankle arthroplasty had a positive effect on the quality of life, daily activities, and motor function recovery of patients with orthopedic ankle injuries. While ankle arthroplasty has potential for clinical application, future high-quality, long-term studies with larger samples and more outcome indicators are warranted to verify these results.

One-Pot Bioelectrocatalytic Conversion of Chemically Inert Hydrocarbons to Imines.

Petroleum hydrocarbons are our major energy source and an important feedstock for the chemical industry. With the exception of combustion, the deep conversion of chemically inert hydrocarbons to more valuable chemicals is of considerable interest. However, two challenges hinder this conversion. One is the regioselective activation of inert carbon-hydrogen (C-H) bonds. The other is designing a pathway to realize this complicated conversion. In response to the two challenges, a multistep bioelectrocatalytic system was developed to realize the one-pot deep conversion from heptane to N-heptylhepan-1-imine under mild conditions. First, in this enzymatic cascade, a bioelectrocatalytic C-H bond oxyfunctionalization step based on alkane hydroxylase (alkB) was applied to regioselectively convert heptane to 1-heptanol. By integrating subsequent alcohol oxidation and bioelectrocatalytic reductive amination steps based on an engineered choline oxidase (AcCO6) and a reductive aminase (NfRedAm), the generated 1-heptanol was successfully converted to N-heptylhepan-1-imine. The electrochemical architecture provided sufficient electrons to drive the bioelectrocatalytic C-H bond oxyfunctionalization and reductive amination steps with neutral red (NR) as electron mediator. The highest concentration of N-heptylhepan-1-imine achieved was 0.67 mM with a Faradaic efficiency of 45% for C-H bond oxyfunctionalization and 70% for reductive amination. Hexane, octane, and ethylbenzene were also successfully converted to the corresponding imines. Via regioselective C-H bond oxyfunctionalization, intermediate oxidation, and reductive amination, the bioelectrocatalytic hydrocarbon deep conversion system successfully realized the challenging conversion from inert hydrocarbons to imines that would have been impossible by using organic synthesis methods and provided a new methodology for the comprehensive conversion and utilization of inert hydrocarbons.

Analyzing mechanisms in Co(i) redox catalysis using a pattern recognition platform.

Redox catalysis has been broadly utilized in electrochemical synthesis due to its kinetic advantages over direct electrolysis. The appropriate choice of redox mediator can avoid electrode passivation and overpotential, which strongly inhibit the efficient activation of substrates in electrolysis. Despite the benefits brought by redox catalysis, establishing the precise nature of substrate activation remains challenging. Herein, we determine that a Co(i) complex bearing two N,N,N-tridentate ligands acts as a competent redox catalyst for the reduction of benzyl bromide substrates. Kinetic studies combining electroanalytical techniques with multivariable linear-regression analysis were conducted, disclosing an outer-sphere electron-transfer mechanism, which occurs in concert with C-Br bond cleavage. Furthermore, we apply a pattern recognition platform to distinguish between mechanisms in the activation of benzyl bromides, found to be dependent on the ligation state of the cobalt(i) center and ligand used.

N-Ammonium Ylide Mediators for Electrochemical C-H Oxidation.

The site-specific oxidation of strong C(sp3)-H bonds is of uncontested utility in organic synthesis. From simplifying access to metabolites and late-stage diversification of lead compounds to truncating retrosynthetic plans, there is a growing need for new reagents and methods for achieving such a transformation in both academic and industrial circles. One main drawback of current chemical reagents is the lack of diversity with regard to structure and reactivity that prevents a combinatorial approach for rapid screening to be employed. In that regard, directed evolution still holds the greatest promise for achieving complex C-H oxidations in a variety of complex settings. Herein we present a rationally designed platform that provides a step toward this challenge using N-ammonium ylides as electrochemically driven oxidants for site-specific, chemoselective C(sp3)-H oxidation. By taking a first-principles approach guided by computation, these new mediators were identified and rapidly expanded into a library using ubiquitous building blocks and trivial synthesis techniques. The ylide-based approach to C-H oxidation exhibits tunable selectivity that is often exclusive to this class of oxidants and can be applied to real-world problems in the agricultural and pharmaceutical sectors.

Atom-Economical Cross-Coupling of Internal and Terminal Alkynes to Access 1,3-Enynes.

Selective carbon-carbon (C-C) bond formation in chemical synthesis generally requires prefunctionalized building blocks. However, the requisite prefunctionalization steps undermine the overall efficiency of synthetic sequences that rely on such reactions, which is particularly problematic in large-scale applications, such as in the commercial production of pharmaceuticals. Herein, we describe a selective and catalytic method for synthesizing 1,3-enynes without prefunctionalized building blocks. In this transformation several classes of unactivated internal acceptor alkynes can be coupled with terminal donor alkynes to deliver 1,3-enynes in a highly regio- and stereoselective manner. The scope of compatible acceptor alkynes includes propargyl alcohols, (homo)propargyl amine derivatives, and (homo)propargyl carboxamides. This method is facilitated by a tailored P,N-ligand that enables regioselective addition and suppresses secondary E/Z-isomerization of the product. The reaction is scalable and can operate effectively with as low as 0.5 mol % catalyst loading. The products are versatile intermediates that can participate in various downstream transformations. We also present preliminary mechanistic experiments that are consistent with a redox-neutral Pd(II) catalytic cycle.

Gold-Catalyzed Formal Hexadehydro-Diels-Alder/Carboalkoxylation Reaction Cascades.

A dual gold-catalyzed hexadehydro-Diels-Alder/carboalkoxylation cascade reaction is reported. In this transformation, the gold catalyst participated in the hexadehydro-Diels-Alder step, switching the mechanism from a radical type to a cationic one, and then the catalyst activated the resulting aryne to form an ortho-Au phenyl cation species, which underwent a carboalkoxylation rearrangement rather than the expected aryne-ene reaction.

A Computer-Interpretable Guideline for COVID-19: Rapid Development and Dissemination.

BACKGROUND: COVID-19 is a global pandemic that is affecting more than 200 countries worldwide. Efficient diagnosis and treatment are crucial to combat the disease. Computer-interpretable guidelines (CIGs) can aid the broad global adoption of evidence-based diagnosis and treatment knowledge. However, currently, no internationally shareable CIG exists. OBJECTIVE: The aim of this study was to establish a rapid CIG development and dissemination approach and apply it to develop a shareable CIG for COVID-19. METHODS: A 6-step rapid CIG development and dissemination approach was designed and applied. Processes, roles, and deliverable artifacts were specified in this approach to eliminate ambiguities during development of the CIG. The Guideline Definition Language (GDL) was used to capture the clinical rules. A CIG for COVID-19 was developed by translating, interpreting, annotating, extracting, and formalizing the Chinese COVID-19 diagnosis and treatment guideline. A prototype application was implemented to validate the CIG. RESULTS: We used 27 archetypes for the COVID-19 guideline. We developed 18 GDL rules to cover the diagnosis and treatment suggestion algorithms in the narrative guideline. The CIG was further translated to object data model and Drools rules to facilitate its use by people who do not employ the non-openEHR archetype. The prototype application validated the correctness of the CIG with a public data set. Both the GDL rules and Drools rules have been disseminated on GitHub. CONCLUSIONS: Our rapid CIG development and dissemination approach accelerated the pace of COVID-19 CIG development. A validated COVID-19 CIG is now available to the public.

Stereodivergent Coupling of 1,3-Dienes with Aldimine Esters Enabled by Synergistic Pd and Cu Catalysis.

Herein we describe the use of synergistic Pd and Cu catalysis for stereodivergent coupling reactions between 1,3-dienes and aldimine esters. By using different enantiomers of the two metal catalysts, all four stereoisomers of the coupling products, which have two vicinal stereocenters, could be accessed with high diastereo- and enantioselectivity. This atom-economical cross-coupling reaction has a wide substrate scope and good functional group tolerance. Our work highlights the power of synergistic catalysis for asymmetric coupling reactions involving Pd-hydride catalysts.

[Molecular mechanism underlying differentiation of HL-60 cells induced by hexamethylene bisacetamide].

The objective of this study was to investigate the effect of hexamethylene bisacetamide (HMBA) on differentiation of HL-60 cells and its possible molecular mechanism. HL-60 cells were co-cultured with different concentrations of HMBA (0.5, 1, 2 mmol/L) for 4 days, then the proliferation was assayed by MTT at different time points. Wright-Giemsa staining was used to observe the change in morphology. Cell differentiation antigen CD11b expression and the altered distribution of cell cycle in HL-60 induced by HMBA were analyzed by flow cytometry. The expressions of c-myc, mad1, p21, p27, hTERT and HDAC1 mRNA were detected by RT-PCR. The results indicated that the proliferation of HL-60 cells was inhibited by HMBA in a time-and-dose-dependent manner. Upon 2 mmol/L HMBA treatment, the HL-60 cells arrested at G(0)/G(1) phase and differentiated into granular line in morphology, with the up-regulation of CD11b expression. The expression of c-myc and hTERT mRNA obviously down-regulated, the expression of p21, p27 and mad1 mRNA up-regulated, while there was no change of the expression of hTERT mRNA. It is concluded that effect of HMBA on the differentiation of HL-60 cells may partly contribute to switch from c-myc to mad1 expression, to up-regulate expressions of p21 and p27 mRNA, and down-regulate hTERT mRNA expression, while there is no relation with the expression of HDAC1 mRNA.

Role of Myc in differentiation and apoptosis in HL60 cells after exposure to arsenic trioxide or all-trans retinoic acid.

Acute promyelocytic leukemia (APL) is highly malignant and frequently expresses the PML-RARalpha (promyelocytic leukemia-retinoic acid receptor-alpha) fusion protein. This fusion protein is targeted by all-trans retinoic acid (ATRA) and arsenic trioxide (As2O3), presently used in APL therapy. We have evaluated effects of ATRA and As2O3 treatment in PML-RARalpha-negative HL60 promyelocytic leukemia cells, harboring amplified c-myc. Characterization of expression and activity of c-Myc and its target genes hTERT (human telomerase reverse transcriptase) and CAD (carbamoyltransferase-dihydroorotase) revealed marked down-regulation in response to ATRA, but not As2O3. We suggest that blockage of terminal differentiation upon As2O3 treatment may be mediated through c-Myc.

Down-regulation of CD44 contributes to the differentiation of HL-60 cells induced by ATRA or HMBA.

CD44 is highly expressed in human acute myeloid leukemia (AML) cells. Some experiments had shown that it was possible to reverse differentiation blockage in AML cells by CD44 ligation with specific antibodies, indicating that CD44 was closely related to the differentiation of leukemia cells. The differentiation of acute promyelocytic leukemia cell line HL-60 cells could be induced by all trans-retinoic acid (ATRA) and hexamethylene bisacetamide (HMBA), but so far the mechanism was not demonstrated clearly. In the present study, we investigated whether ATRA or HMBA induced the growth arrest of HL-60 cells by down-regulating the expression of CD44. The results indicated that the proliferation of HL-60 cells was obviously inhibited and the differentiation was induced by both ATRA and HMBA. The decreased expression of CD44 and cyclin E mRNA, and the increased expression of p27 and p21 at mRNA levels were observed. Furthermore, there was a negative correlation between the expression of CD44 and p27. It was concluded that ATRA and HMBA played a role in the differentiation induction of HL-60 cells, which was mediated by the down-regulation of CD44, accompanied by down-regulation of cyclin E, and up-regulation of p27 and p21 mRNA.

Down-regulation of TRRAP-dependent hTERT and TRRAP-independent CAD activation by Myc/Max contributes to the differentiation of HL60 cells after exposure to DMSO.

Myc/Max/Mad often play pivotal roles in the proliferation, apoptosis, differentiation and cell cycle progress of leukemia cells. Myc and Mad are known to be unstable proteins and their expression is tightly regulated throughout cell cycle progression and differentiation. Usually, c-Myc expression is implicated in cell growth and proliferation, and the deregulated expression of c-Myc in both myeloid leukemia cells and normal myeloid cells not only blocks terminal differentiation but also its associated growth arrest. HL60 cells could be induced to differentiate into mature granulocytes by DMSO in vitro, but the mechanism of this effect has not been elucidated clearly. We proposed the hypothesis that down-regulation of c-Myc expression by DMSO contributed to the differentiation of HL60 cells by way of activating target genes hTert and CAD. The results showed that c-Myc expression was down-regulated in differentiated HL60 cells but not in exponentially-growing HL60 cells, without or with the target gene activation of hTert and CAD, respectively. Further study indicated that hTert activation is TRRAP-dependent while CAD activation is TRRAP-independent. On the other hand, up-regulation of P(21) and P(27) and down-regulation of cyclinA and cyclinE also play important roles in induction of the terminal differentiation of HL60 cells. Our results support the hypothesis that c-Myc expression and activation of target genes for hTert and CAD play critical roles in the proliferation of HL60 cells, while down-regulation of c-Myc expression and activation of target genes for hTert and CAD contributed to the terminal differentiation of HL60 cells after exposure to DMSO in vitro.

Immune tolerance in pancreatic islet xenotransplantation.

AIM: To observe the effect of tail vein injection with donor hepatocytes and/or splenocytes on the islet xenotransplantation rejection. METHODS: New-born male pigs and BALB/C mice were selected as donors and recipients respectively. Islet xenotransplantation was performed in recipients just after the third time of tail vein injection with donor hepatocytes and/or splenocytes. Macrophage phagocytosis, NK(natural killing cell) killing activity, T lymphocyte transforming function of spleen cells, antibody forming function of B lymphocytes, and T lymphocyte subsets were taken to monitor transplantation rejection. The effects of this kind of transplantation were indicated as variation of blood glucose and survival days of recipients. RESULTS: The results showed that streptozotocin (STZ) could induce diabetes mellitus models of mice. The pre-injection of donor hepatocytes, splenocytes or their mixture by tail vein injection was effective in preventing donor islet transplantation from rejection, which was demonstrated by the above-mentioned immunological marks. Each group of transplantation could decrease blood glucose in recipients and increase survival days. Pre-injection of mixture of donor hepatocytes and splenocytes was more effective in preventing rejection as compared with that of donor hepatocyte or splenocyte pre-injection respectively. CONCLUSION: Pre-injection of donor hepatocytes, splenocytes or their mixture before donor islet transplantation is a good way in preventing rejection.

Effect of arsenic trioxide on cytokine expression by acute promyelocytic leukemia cells.

OBJECTIVE: To detect the expression of cytokines by acute promyelocytic leukemia (APL) cells before and after exposure to arsenic trioxide. METHODS: Diagnoses were performed according to the FAB cytological classification criteria and cytogenetic criteria. Bone marrow or blood samples from APL patients were collected in heparinized tubes, then primary APL cells were separated by traditional Ficoll-Hypaque density centrifugation and purified after adherence to plastic surfaces. IL-1(beta), IL-6, IL-8, TNF alpha and G-CSF levels in the leukemia cell culture supernatants were detected by ELISA. At the same time, nitro blue tetrazolium (NBT) reduction test was used to detect the differentiation of APL cells. RESULTS: After 96 hours exposure to arsenic trioxide, 10 - 6 mol/L in vitro or 10 mg/d in vivo, APL cells showed a significant increase of IL-1(beta) (P < 0.05) and G-CSF (P < 0.05) production, and a significant decrease of IL-6 (P < 0.05) and IL-8 (P < 0.05). However, there was no obvious variation of TNF alpha when compared with APL cells without exposure to arsenic trioxide. On the other hand, the proliferation ratio of APL cells in vitro was statistically correlated to the IL-1(beta) secretion ratio or G-CSF secretion ratio. The cell number ratio in patients with detectable IL-1(beta) or G-CSF was higher than that without detectable IL-1(beta) or G-CSF. CONCLUSION: IL-1(beta) and G-CSF secretion may play an important role in the proliferation of APL cells after exposure to arsenic trioxide.

The role of donor hepatocytes and/or splenocytes pre-injection in reducing islet xenotransplantation rejection.

OBJECTIVE: To observe the effect of tail vein injection with donor hepatocyte and/or splenocyte on islets xenotransplantation rejection. METHODS: New-born male pigs and BALB/C mice were selected as donors and recipients respectively. Islets xenotransplantation was performed in recipients just after the third time of tail vein injection with donor hepatocytes and/or splenocytes. Macrophage phagocytosis, NK killing activity, T lymphocyte transforming function of spleen cells, antibody forming function of B lymphocytes, and T lymphocyte subsets were taken to monitor transplantation rejection. The effects of this kind of transplantation were indicated as variation of blood glucose and survival days of recipients. RESULTS: Streptozotocin (STZ) succeeded in inducing diabetes mellitus models of mice. Pre-injection of donor hepatocytes, and splenocytes or their mixture via tail vein was effective in preventing donor islets transplantation from rejection, which was demonstrated by the mentioned immunological marks. And each group of transplantation could decrease the blood glucose of recipients and prolong the survival days. Pre-injection of mixture of donor hepatocytes and splenocytes was more effective in preventing rejection than pre-injection of donor hepatocytes or splenocytes separately. CONCLUSION: We propose that pre-injection of donor hepatocytes, splenocytes separately or their mixture before donor islets transplantation is a good way to prevent rejection.