Room-temperature electrochemical hydroxyalkylation of pyridines

Domain: chemistry
Published: 2026-04-13
Editorial outcome: major_revision

Abstract
Direct and selective functionalization of ubiquitous pyridine scaffold is of paramount importance across numerous fields, yet remains challenging due to the inherent inertness of pyridines and the presence of competing reactive sites. Here, using an undivided cell equipped with a zinc cathode, a graphite anode, and a TEMPO mediator, we report a room-temperature electrochemical strategy that enables the direct hydroxyalkylation of diverse pyridines with readily available carbonyl compounds. This method features a broad substrate scope, operational simplicity, metal-free conditions, and high step/atom economy. It overcomes the limitations of conventional approaches requiring pre-functionalized substrates or stoichiometric activators, establishing a practical platform for direct access to C4- or C2-hydroxyalkyl pyridines, depending on the substitution pattern of the pyridine substrates. Mechanistic studies reveal that acetic acid activates both reactants, and the products are formed via cross-coupling of cathodic reduction-induced pyridyl radicals and hydroxy α-radicals followed by protonation and anodic oxidative aromatization.

Introduction
Pyridine represents most prevalent N-heterocyclic scaffold in FDA-approved pharmaceuticals [1] and serves as a key structural motif in numerous agrochemicals, functional materials, ligands, and natural products.[2-6] Among its derivatives, 4-hydroxyalkylated pyridines constitute a significantly important subclass due to their distinctive properties and broad applications.[7-14] As illustrated in Fig. 1A, representative examples include pyridoxine (1, a natural form of vitamin B6),[11] a CCR6 antagonist (2) for autoimmune diseases,[7] and LY2874455 (3), a commercially available second-generation FGFR inhibitor used in cancer therapy.[8] Additionally, compounds 4 and 5 function as a surface-grafting agent for MOF-based polymers and a precursor for polymer-supported catalysts, respectively.[12,13] Despite their significance, synthetic routes to 4-hydroxyalkyl pyridines remain notably scarce. Conventional syntheses typically rely on either initial preparation of C4-halopyridines followed by lithiation, nucleophilic addition, and protonation or Grignard addition to preinstalled 4-pyridyl ketones and subsequent hydrolysis (Fig. 1B).[15,16] These methods often require harsh conditions, exhibit poor functional group tolerance, and suffer from low step and atom economy, and generate substantial metal wastes. Thus, the development of efficient and direct strategies for diverse synthesis of 4-hydroxyalkyl pyridines is desirable.

Despite the challenges posed by pyridine's inherent thermodynamic stability, kinetic inertness, and multiple competitive reaction sites, several elegant strategies have been established for direct functionalization at the C2[17-22] and C3[23-32] positions. In contrast, functionalization at the C4 position typically relies on multistep sequences employing pre-activated substrates such as pyridinium salts, N-oxides, or pyridyl salts (Fig. 1C-1, 1C-2 and 1C-3).[33-47] Although Minisci-type radical addition enables direct functionalization of heteroarenes,[17-22] it generally yields C2-substituted products alongside minor amounts of C4 regioisomers (Fig. 1C-4). In this context, cross-coupling between pyridyl radicals generated via single-electron reduction (SER) and polarity-matched radicals represents a promising strategy for achieving direct C4 functionalization, particularly for C2-substituted pyridines. Nonetheless, this objective remains a yet underdeveloped subject in synthetic chemistry.

In recent years, electrochemistry has emerged as an appealing tool in synthetic chemistry owing to the facile tunability of key reaction parameters (e.g., electrolyte, electrode, current, and potential).[48-72] Capitalizing on these advantages, we previously reported a direct method for constructing N-heterocyclic vicinal amino alcohols via electroreductive cross-coupling of the quinolyl C2-site with carbonyl compounds.[73] Inspired by this work, we envisioned a practical electrochemical cross-coupling strategy for direct C4-functionalization of C2-substituted pyridines (A) with carbonyls (B). As shown in Fig. 1D, our proposed strategy involves the simultaneous cathodic reduction of pyridine A and carbonyl agent B, generating radicals A-1 and B-1. Subsequent radical-radical cross-coupling, followed by protonation and anodic oxidative aromatization of the resulting intermediate C-1, affords the desired product C. Notably, being significantly different from the reactive quinoline reactions employing a sacrificial zinc anode,[73] the electrons released from the anodic oxidation of the coupling adduct (C-1) in this approach are subsequently consumed in the cathodic reduction process. Such an electrochemical strategy[74-76] substantially improves the synthetic efficiency while avoiding the need for sacrificial oxidants or reductants, thereby providing a greener and more environmentally benign route to functionalize pyridines.

However, realizing the above synthetic purpose presents several critical challenges: (1) Rate matching issue: The high reduction potentials (E < -2.2 V vs. SCE) of pyridines need to be lowered to match those of carbonyl compounds. This adjustment is essential for compatible reaction kinetics and to prevent overreduction of the more reactive carbonyl agents (B) into alcohols (B''). (2) Selectivity control: Diverse radical intermediates generated during pyridine reduction require precise reaction modulation for high C4-regioselectivity (C rather than C'). Additionally, radicals derived from carbonyl reduction should exhibit sufficient persistence to timely trap the pyridyl radicals in situ. This persistence is crucial for achieving high chemoselectivity by suppressing homocoupling processes (A' and B'). (3) Requiring effective redox neutral electrolysis: Coupling adduct (C-1) must rapidly diffuse away from the cathode's electrical double layer to undergo efficient anodic oxidative aromatization, thereby preventing its decomposition to achieve efficient transformation.

Here, by integrating acetic acid activation of both reactants with TEMPO-mediated oxidative aromatization, we report a room-temperature electrochemical strategy for direct C4- or C2-hydroxyalkylation of pyridines with readily available carbonyls. This method circumvents the limitations of conventional pyridyl functionalization that require pre-functionalized steps or stoichiometric activators. Furthermore, it achieves air/moisture tolerance, broad substrate scope, operational simplicity, metal-free conditions, high step and atom economy, and diverse synthesis.

Results
Reaction condition optimizations
To evaluate the feasibility of the envisioned synthetic protocol, we chose the coupling reaction between 2-phenylpyridine (A ) and 4'-methoxyacetophenone (B ) as a model system for parameter screening. The reaction was conducted in an undivided cell equipped with a graphite anode and a zinc cathode, using N,N-dimethylformamide (DMF) as the solvent and tetra-n-butylammonium tetrafluoroborate (n-Bu₄NBF₄, 0.1 M) as the electrolyte. The reaction was applied with constant current of 25 mA for 9 h in the presence of TEMPO (2,2,6,6-tetramethylpiperidin-1-oxyl) and acetic acid (AcOH). As shown in Table 1, this reaction afforded the C4-hydroxyalkyl product C with an optimal isolated yield of 81% (entry 1). Excellent regioselectivity was confirmed by GC-MS and HPLC analyses (C :C ' > 20:1). Control experiments confirmed that both electricity (entry 2) and AcOH (entry 3) were indispensable for the reaction. The absence of TEMPO significantly decreased the product yield, along with low conversion of B (entry 4), indicating that TEMPO acts as a reaction mediator. Current variations proved critical: no reaction was occurred at 15 mA (entry 5) with substrates fully recovered, while increasing the current to 30 mA reduced the yield (entry 6). Moderate yields were obtained when adjusted the stoichiometric ratio of TEMPO/AcOH or replaced AcOH with HCOOH (entries 7–9). Neither ferrocene nor n-Bu NI enabled the formation of C1 (entries 10–11). This is attributed to the insufficient oxidizing capability of ferrocenium ions and elemental iodine, which are unable to effectively promote the oxidation of the intermediate—a key step in product formation. Additionally, these mediators may undergo competitive side reactions, further reducing the overall reaction efficiency. Solvent screening demonstrated that DMF outperformed NMP and acetone (entry 12). Cathode substitution (graphite or nickel) diminished the yield (entry 13), while platinum or zinc anodes performed the reaction poorly (entry 14). Notably, identical yields were obtained under argon and air atmospheres (entry 15), whereas the addition of H O (10 equiv.) led to a slight decrease in yield (entry 16). This indicates that the reaction exhibits good compatibility with water and oxygen. Moreover, both extended and shortened reaction times failed to improve the outcomes (Supplementary Table S1).

Substrates scope investigation
With the optimal conditions established, we set out to evaluate the substrate scope of the developed synthetic protocol. As shown in Fig. 2, diverse ketones and aldehydes (B –B , B ; see Supplementary Fig. S1 for their structures) were employed to couple with pyridines. Ketones lacking redox-sensitive groups (B –B ) gave C4-hydroxyalkyl pyridines (C –C ) in moderate to good isolated yields. Electron-donating groups and halogens were well tolerated (C –C ), 1-(4-chlorophenyl)ethan-1-one (B ) required shorter reaction time to prevent hydrodehalogenation of product C . After stoichiometric adjustment, trifluoromethyl-containing aryl ketone B was able to afford product C in moderate yield. 3-Acetylthiophene provided product C in acceptable yield. Cyclic and different alkyl-substituted ketones reacted smoothly (C –C ). Trifluoromethyl (B –B ) and vinyl ketones (B ) also generate the desired products in moderate yields. Noteworthy, 3-penten-2-one yielded C along with the regioisomer C , the latter suggests a spin center shift (SCS) of the hydroxy α-radical. Gratifyingly, aryl aldehydes (B –B ) also underwent efficient transformation, delivering products C –C in reasonable yields. Notably, cyclohexanone (B ) failed to react productively, and pyridine homocoupling product was obtained instead, which stems from the excessively high reduction potential of cyclohexanone.

Next, we investigated the reaction of diverse N-heteroarenes (A −A , Supplementary Fig. S1). Given the obvious variation in reduction potentials among these substrates, adjustments to the standard reaction conditions were necessary (Fig. 3). A range of functionalities (e.g., −OMe, −Cl, −i-Pr, −CO Et, −Me, −TMS, −F) proved compatible, affording the desired products (C −C ) in moderate to good yields. Notably, electron-rich pyridines required either a graphitic cathode or extended reaction time to improve the yields. Thiophene-substituted pyridines yielded the desired products (C and C ) within shorter times. Electron-withdrawing groups (−CO Et, −CF ) and unsaturated substituents (vinyl, alkynyl) were well tolerated (C −C ). It is noteworthy that 2,2'-bipyridine underwent 4,4'-bisfunctionalization to afford compound C , demonstrating the potential of the present chemistry in developing valuable bidentate N-ligands. Pyridine (A ) and 2-picoline (A ) also underwent hydroxyalkylation efficiently, affording the corresponding C4-functionalized products (C and C ) in 40% and 24% yields, respectively. Interestingly, when equimolar amounts of aldehyde and amine were added, the in-situ-generated imines (B −B ) could also be used in the coupling reaction with A , affording the C4-aminoalkyl pyridines (C , C ) in reasonable yields. This example further demonstrates the broader potential of the present synthetic method. In addition to the C4-functionalization of C2-substituted pyridines, the method is also applicable to C3- and C4-substitued pyridines to afford preferable C2-functionalized products. Interestingly, extending the reaction time for the coupling of 4-phenylpyridine (A ) and B from 4 h to 9 h shifted the product from the C2-hydroxyalkyl species (C ', 41%) to the 2,6-dihydroxyalkyl product (C '). 4-(Trifluoromethyl)pyridine (A ) also reacted smoothly with 4-methoxyacetophenone, affording the desired product (C ') in 29% yield. Both 3-phenylpyridine (A ) and 3-(trifluoromethyl)pyridine (A ) underwent smooth transformation to form their respective C2-functionalized products C ' and C ', with this selectivity presumably governed by steric effects. In contrast, the disubstituted pyridines (A , A ) afforded products C and C with exclusive C4-selectivity. Fused pyridines were also suitable substrates. For instance, thieno[3,2-b]pyridine (A ) and furo[3,2-b]pyridine (A ) afforded mixtures of major C4- and minor C2-regioisomers (C /C ', C /C '). Conversely, quinolines (A , A ) and 4,7-phenanthroline (A ) selectively provided C2-functionalized products (C '−C '), a class of potential hemilabile bidentate ligands. These examples show that the present synthetic protocol is also applicable for direct functionalization of fused pyridines, and the regioselectivity differences are attributed to the combined effects of radical stability and the steric influence.

Investigations of the reaction mechanism
To gain mechanistic insights into the established reaction, we initially conducted various control experiments. As shown in Fig. 4A, separate treatment of 2-phenylpyridine A and 4-methoxyacetophenone B under the standard conditions yielded the homocoupling products D and D , respectively (Fig. 4A-I and 4A-II). Introducing the radical scavenger TEMPO (2.5 equiv.) into the model reaction completely suppressed the formation of product C ; instead, a TEMPO-trapping pyridine adduct TEMPO-B was observed (Fig. 4A-III). These results indicate a radical-involved pathway. Subsequently, a radical clock experiment using cyclopropyl ketone B and A afforded a ring-opening product (C ), demonstrating that the ketone also acts as a radical precursor (Fig. 4B). Finally, a kinetic isotope effect (KIE) experiment with equimolar A and A -d afforded the desired product in 34% yield with a k /k value of 1.94 (Fig. 4C). This result suggests that C–H bond cleavage at the pyridyl C4 position is likely involved in the rate-determining step, though the presence of other rate-limiting processes cannot be excluded.[77]

Next, we performed cyclic voltammetry (CV) and open-circuit voltage (OCV) experiments to probe the electrochemical behavior of the model reaction (Fig. 4D and Supplementary Fig. S9). The CV of 2-phenylpyridine (A ) displayed a reversible redox process with a reduction peak at −2.42 V (vs. SCE in DMF), while 4′-methoxyacetophenone (B ) exhibited a reduction potential of −2.28 V. Upon addition of AcOH, both A and B showed two reduction peaks (A : −2.26 V and −2.79 V; B : −2.13 V and −2.55 V). The positive shift in reduction potentials and the enhanced catalytic current show that AcOH activates both reactants. Their similar onset potentials (−1.68 V) and peak potentials provide compelling evidence for concurrent reduction. This conclusion was further supported by OCV experiments (Supplementary Fig. S9), which revealed a stable cathodic plateau at −1.95 V (vs. SCE). Notably, the oxidation peak intensities of A and B decreased in the presence of AcOH, suggesting that AcOH stabilizes their radical intermediates generated by cathodic reduction.

Based on the above findings and the transformations employing redox neutral electrolysis,[78,79] we propose a plausible reaction pathway for the reaction of A and B to form product C (Fig. 4E). Initially, cathodic reduction of proton-activated 4'-methoxyacetophenone (B -H) generates a persistent benzyl radical (or hydroxy α-radical B '), while electroreduction of protonated 2-phenylpyridine (A -H) yields the pyridyl radical INT-1. Simultaneously, anodic single-electron oxidation of TEMPO (I) forms oxoammonium species II. Subsequent cross-coupling of radicals INT-1 and B ' affords the coupling adduct INT-2. Species II then mediates oxidative aromatization of INT-2, delivering product C along with release of TEMPOH (III). Finally, anodic oxidation of III regenerates TEMPO (I), completing the catalytic cycle. Calculation of the Faradaic efficiency (FE) for this transformation revealed a relatively low value under standard conditions (3.86%; Supplementary Table S1). Given that hydrogen evolution is not significant for most substrates, we attribute this low FE primarily to an unproductive cycle, wherein radicals generated upon single-electron reduction of the substrate are re-oxidized by TEMPO prior to cross-coupling. As product formation necessitates radical cross-coupling following simultaneous single-electron reduction of substrates with high reduction potentials, the observed low FE remains acceptable for reaction conducted in an undivided cell with non-sacrificial anode. Comparable FE values have been reported in similar electrochemical systems involving multiple single-electron transfers of the substrates with high reduction potential.[80]

Further, DFT calculations were applied to analyze the proposed reaction pathway. All geometries were fully optimized, and the parameters are given in Supplementary Table S3 in the SI. As shown in Fig. 5, A -H is initially generated from the protonation of A due to the presence of acetic acid in the reaction system. This protonation step will facilitate a subsequent single electron transfer process and generate radical INT-1. Spin density analysis of INT-1 reveals that the electron spin predominantly located on the C4 and C6 positions, with little spin density at C2 position. Mulliken spin population calculation further supports that the electron population on C4 position (0.367) is significantly higher than on C2 position (0.0896). These results suggest that the C4 position will be more reactive towards radical coupling. To study the chemoselectivity of the radical coupling step, the reaction energy barrier between radical B ' (Generated from cathodic reduction of ketone B ) and INT-1 are calculated for C2 and C4 position. The radical combination energy barrier at C4 position is 9.7 kcal/mol, which is 2.3 kcal/mol lower than that at C2 position. Thus, the radical coupling process will preferentially undergo at C4 position and afford INT-2 rather than INT-3. Subsequently, given the nucleophilic positions in INT-2 and potential hydrogen donor characteristics, the subsequent reaction between TEMPO+ (Generated from the anodic oxidation of TEMPO) may have multiple pathways. When the C3 carbon attack the nitrogen in the N=O double bond of TEMPO+, the reaction energy barrier is 21.1 kcal/mol, which is relatively high. In addition, this step is also thermodynamically unfavorable (ΔG=17.7 kcal/mol). Thus, the following cope elimination from INT-5 to aromatization and forming INT-4 is unlikely to occur. Attempts to locate a stable intermediate via nucleophilic attack from the nitrogen atom of INT-2 were unsuccessful, possibly due to the steric hinderance between the tera-methyl groups in TEMPO+ and the phenyl group in INT-2. Alternatively, a both kinetically and thermodynamically favorable hydride transfer mechanism was identified,[81-83] as 1,4-dihyropyridines are well-documented hydrogen donors.[84-86] The hydrogen transfer between INT-2 and TEMPO+ has a low energy barrier (15.9 kcal/mol) and favored thermodynamics (ΔG = -37.1 kcal/mol), which will lead to the formation of protonated product INT-4 and TEMPOH. Final deprotonation mediated by acetate affords the desired product C . This deprotonation step may also be assisted by substrate A .

Derivatizations of the products
Finally, we explored the synthetic utility of the developed methodology. Even with reduced loadings of TEMPO/AcOH (Fig. 6A), the reaction of A (3 mmol) with B (9 mmol) afforded the product C in good yield (62%, 0.57 g). This yield represents a significant improvement over the multistep procedure reported previously[14] and demonstrates the scalability potential of the protocol. As shown in Fig. 6B, treating C to HCl in toluene (12 h) followed by neutralization afforded the 1,1-diarylethylene D in 85% yield. Hydroboration-oxidation of D gave the secondary alcohol D in 88% yield, while epoxidation with m-CPBA followed by ring-opening yielded the vicinal diol D in 50% yield. Enabled by the pyridyl directing group, palladium-catalyzed ortho-C–H acylation of the phenyl moiety in C provided D in 39% yield. Direct esterification of C with ibuprofen furnished D in 51% yield. We next applied the synthetic method to the functionalization of bioactive substrates (Fig. 6C). Reaction of Isophorone (B ) with A gave the desired product C in merely 16% yield, while the 1,3-rearrangement product C predominated with a 56% yield. Tonalide (B ), Epoxyprogesterone (B ) and L-menthol-derived ketone (B ) reacted efficiently with A to exclusively afford C4-functionalized pyridines (C –C ). Furthermore, the reactions of Myrtenol-derived pyridine A and Abametapir A with B proceeded smoothly, affording C and C in moderate to good yields.

Methods
General procedure for the hydroxyalkylation of pyridines in DMF
An undivided electrolytic cell equipped with a magnetic stir bar was subsequently charged with TEMPO (15.6 mg, 0.1 mmol, 0.5 equiv.), A (0.2 mmol, 1 equiv.), carbonyl substrate B (0.6 mmol, 3 equiv.), n-Bu NBF (330 mg, 0.1 M), DMF (10 mL) and AcOH (58 uL, 1 mmol, 5 equiv.) under air condition. After fully stirring and dissolution, a graphite plate anode and a zinc plate cathode were installed. Then the cell was electrolyzed at room temperature under a constant current of 25 mA for 9 hours (If not otherwise mentioned). Upon the completion of electrolysis, the resulting mixture was poured into saturated NH Cl solution (50 mL) and the electrodes and cell were rinsed with EtOAc, then extracted the mixture with EtOAc (35 mL × 3). The combined organic layers were dried over anhydrous Na SO , filtered, and concentrated under reduced pressure. The resultant residue was purified by preparative TLC on silica to afford hydroxyalkylated pyridines C.
(Note: Both the dimensions of graphite and zinc electro des are approximately 5.25 cm × 0.8 cm × 0.2 cm. The effective immersion areas of graphite anode and zinc cathode are approximately 2.56 cm2.)

Peer review rounds
[{"round": 1, "reviewer_comments": {"reviewer_1": "This manuscript presents a novel and elegant electrochemical strategy for the direct C4-hydroxyalkylation of pyridines using\nreadily available carbonyl compounds. The work addresses a significant synthetic challenge—selective C4 functionalization\nof pyridines, featuring operational simplicity, transition metal-free conditions, and excellent air/moisture tolerance. The\nauthors convincingly demonstrate broad substrate scope, scalability, and mechanistic insight through complementary\nexperimental and computational studies. By cleverly employing paired electrolysis, the method achieves redox-neutral\ntransformation with high step and atom economy. Overall, this is a well-executed and impactful study, I recommend its\npublication in Nature Communications after the following minor points are addressed.\n(1) Although TEMPO has been demonstrated to be an effective mediator in the oxidative aromatization step, the authors\nshould provide a brief explanation in the “Reaction Condition Optimization” section as to why alternative mediators, such as\nferrocene, failed to facilitate the transformation.\n(2) The failure of cyclohexanone (B40) is noted, but no explanation is provided. Is this due to sterics, reduction potential, or\nradical stability? A brief comment would strengthen the scope discussion.\n(3) For aldehydes, yields are generally moderate. Are there competing side reactions (e.g., over-reduction to alcohols or\nhomo-coupling to vicinal diols)?\n(4) The switch in regioselectivity for quinolines and phenanthroline (C2 vs. C4) is attributed to sterics/radical stability?\n(5) For the gram-scale reaction, were any adjustments (e.g., electrode size, stirring rate) necessary?\n(6) The abbreviation “SCS” (spin center shift) should be defined upon first use.\n(7) In the introduction, references to “Scheme 1c1–1c3” should be checked for accuracy (currently labeled as 1c1, 1c2, etc.).\n(8) The phrase “ubiquitous pyridine rings” could be rephrased to “ubiquitous pyridine scaffold” for precision.\n(9) Check formatting of references (e.g., reference 8e is missing, and 8f appears as 8g).", "reviewer_2": "The manuscript describes a TEMPO-mediated paired electrolysis that delivers C4 hydroxyalkylated pyridines from ketones\nand aldehydes at room temperature, with CV, control experiments, radical clocks, KIE, and DFT used to support a\nmechanism involving concurrent formation of pyridyl and hydroxy α radicals followed by oxidative aromatization. The overall\nconcept—direct C4 functionalization of pyridines with simple carbonyls under formally redox neutral conditions in an\nundivided cell—is attractive and addresses a relevant synthetic problem, although the extent of conceptual advance may be\nsomewhat overstated.\n1. C4 selectivity and “inert pyridines”\nThe presentation of C4 selectivity and “inert pyridines” raises major concerns. Most of the high performing examples are 2\naryl or otherwise 2 substituted pyridines, where both the DFT analysis and simple qualitative reasoning indicate that spin\ndensity is biased toward C4/C6; accordingly, the observed C4 functionalization appears to be largely dictated by substrate\nelectronics rather than by a generally applicable site selectivity control element. The only example using unsubstituted\npyridine (C46) requires substantially modified conditions (graphite cathode, 35 mA, pyridine in excess) and still gives only\n40% yield with 10:1 C4:C2, suggesting that intrinsic C4 control on a simple pyridine is modest and sensitive to conditions.\nSeveral substrates also give regioisomeric mixtures or even C2 preference (e.g., fused systems C54/C54′, C55/C55′),\nindicating that the method is not generally C4 selective across unbiased pyridines. This contrasts with the title and abstract\n(“direct C4 hydroxyalkylation of inert pyridines”, “broad substrate scope”, “C4 selective”), which imply a more general and\nelectronics agnostic platform. It would therefore be helpful either to moderate these claims or to broaden the scope (e.g., to\nsimple 3 and 4 substituted and more diverse mono substituted pyridines) and provide full regioisomer profiles to support a\nclaim of general C4 selectivity.\n2. Electrochemical concept: paired vs linear paired electrolysis\nThe electrochemical design does qualify as paired electrolysis in the usual synthetic sense: pyridine and the carbonyl\ncompound are reduced at the cathode, while TEMPO is oxidized at the anode to TEMPO+, which then oxidizes the\ndihydropyridine intermediate to the aromatic product, with TEMPO/TEMPOH being regenerated so that both electrodes are\nused productively and no sacrificial anode or stoichiometric oxidant is required.\nHowever, the manuscript also uses the more specific term “linear paired electrolysis”, and this particular designation is not\nclearly supported by the data. The discussion attributes “linear” behavior to diffusion of the coupling adduct away from the\ncathodic double layer prior to anodic oxidation, but the evidence for a dominant linear redox cycle remains qualitative:\nthere is no quantitative demonstration that a single redox couple is continuously shuttling between electrodes or that the majority\nof current is carried by such a cycle (e.g., via faradaic efficiency, partitioning of anodic vs cathodic processes, potential\nprofiles, or mediator mass balance). In this context, describing the system simply as “paired” or “redox neutral TEMPO\nmediated electrolysis” would seem more appropriate than emphasizing the stricter, mechanistically defined “linear paired”\nlabel.\n3. Missing quantitative electrochemical characterization\nGiven how central “paired” and “redox neutral” electrolysis are to the narrative, the lack of faradaic efficiency and current\nefficiency data represents an important gap. It would be informative to see faradaic efficiencies for a few key examples\n(ideally including the gram scale experiment) to support the redox neutral description and to indicate how much of the\ncharge is directed into product formation rather than side processes such as overreduction, solvent/electrolyte oxidation, or\nunproductive mediator turnover, especially in the context of strong claims about efficient paired electrolysis.\n4. Conceptual claims on “new reactivity paradigms”\nThe conclusion’s statement that merging reductive dearomatization with radical transformation “unlocks new reactivity\nparadigms for inert chemical systems” appears stronger than warranted by the current data set. Related “dearomatize then\nfunctionalize/oxidize” strategies are already known, for example in phosphonium/Zincke pyridinium chemistry,\ndihydropyridine hydride transfer, and other radical/polar dearomatization chemistry, and in this study the concept is mainly\ndemonstrated for electronically activated 2 substituted pyridines with limited exploration of unbiased pyridines or other “inert”\nsubstrates. A more measured formulation (for example, indicating that the work “illustrates a strategy that may be extendable\nto other systems”) would better reflect what has been established experimentally.\n5. Supporting Information (NMR spectra)\nAs a minor point regarding the Supporting Information, the ¹H NMR spectra would benefit from expanded views of the\naromatic region; at present these regions are quite compressed, making it difficult to clearly assess coupling patterns and\nindependently verify the assigned substitution patterns.\n6. Overall assessment\nIn summary, the transformation is synthetically useful, and the general electrochemical strategy is of interest, but several\ncentral claims—general C4 selectivity across “inert” pyridines, the “linear paired electrolysis” designation, and the\ndescription of “new reactivity paradigms”—are not fully supported by the current scope and data. The electrochemical\ncharacterization remains incomplete (no faradaic efficiencies and limited electroanalysis), and the substrate scope is heavily\nweighted toward electronically biased 2 substituted pyridines, with only modest performance on truly unbiased pyridines.\nTaken together, these points suggest that, in its present form, the work does not yet reach the level of conceptual generality\nand mechanistic support expected for Nature Communications, and may ultimately be better aligned with a more specialized\njournal once the claims and presentation are adjusted accordingly.", "reviewer_3": "In this manuscript, Zhang and co-workers report an electroreductive C–C bond formation between pyridine derivatives and\ncarbonyl compounds. By leveraging paired electrolysis, the authors achieve direct C4-functionalization of pyridines under\nmild acidic conditions without the need for pre-activation. This transformation employs operationally practical, air- and\nmoisture-tolerant conditions, representing an innovative and impactful example of electrochemical radical–radical\ncoupling.\nThe manuscript is well-structured and clearly written. The electrochemical protocols are described in detail and have been\nsuccessfully applied to the late-stage functionalization of complex bioactive molecules, underscoring the practical utility of\nthe method. Given the novelty, methodological rigor, and scientific significance of this work, I recommend the manuscript for\npublication in Nature Communications following the minor revisions listed below.\n1) What exactly happened when acetone was used as the solvent, were any byproducts observed?\n2) In this work, the reaction is achieved by employing TEMPO as a redox mediator. Can the reaction be achieved in high\nyield through a simple workup after completing the electrolysis with the condition employing a zinc anode?\n3) Reaction times are reported in the optimization of reaction conditions. However, faradaic equivalents (F/mol) provide a\nmore informative measure of efficiency than reaction time, enabling a marked improvement in the reaction's reproducibility.\n4) Was the excess acetophenone fully consumed and converted into by-products during the reaction?\n5) In terms of the mechanism, it is very challenging to have two different radicals to combine due to the low concentration\nand short lifetime. The DFT calculation further illustrates the feasibility of the mechanism. Can the authors explain why the\nalkyl ketones do not proceed smoothly?", "reviewer_4": "In this work, Zhang and coworkers developed an innovative acetic acid-activated electrolysis strategy for direct C4-\nfunctionalization of inert pyridines with ketones/imines at room temperature. This method enables the introduction of\nelectrophiles into pyridine scaffolds without pre-functionalization steps, demonstrating good functional group tolerance and\nbroad synthetic utility. Mechanistic investigations, including experimental studies and DFT calculations, provide insights into\nthe reaction mechanism and chemoselectivity. Overall, this work presents an efficient electrochemical approach for C4-\nfunctionalization of inert pyridines; however, several issues require clarification before further consideration.\n1. Table 1 shows TEMPO as a reaction component, while Figure 1A-III indicates TEMPO inhibits product formation. This\napparent contradiction requires explanation.\n2. In Figure 1A, the authors mentioned that homocoupling occurs when A1 or B1 is reacted individually. However, under\nstandard conditions where both substrates are present simultaneously, heterocoupling is exclusively favored with no\ndetectable homocoupling products. This selectivity pattern requires mechanistic elucidation.The authors are encouraged to\nperform computational studies to provide a reasonable explanation.\n3. The KIE in Figure 1C is interpreted as evidence for rate-determining C–H cleavage. However, following established\nliterature (Angew. Chem. Int. Ed. 2012, 51, 3066), KIE data alone may not definitively identify the RDS. The text should be\nrevised to acknowledge alternative interpretations.\n4. Please clarify: (1) Is \"M06-2x-dd3\" in Figure 2 a typo? (2) Why were different solvation models (PCM for optimization,\nSMD for energies) employed? Reference support for this approach should be provided.\n5. The authors claim “the electron spin predominantly located on the C4 and C6 positions, with little spin density at the C2\nposition.” However, only the spin populations at C2 and C4 are labeled in Figure 1. It is suggested to label the spin\npopulations at N and C6 as well.\n6. Figure S10D is identical to Figure 1. Please remove this redundant figure."}, "author_response": "Response to Reviewers\n\nREVIEWER COMMENTS\nReviewer #1 (Remarks to the Author):\nThis manuscript presents a novel and elegant electrochemical strategy for the direct C4-\nhydroxyalkylation of pyridines using readily available carbonyl compounds. The work addresses a\nsignificant synthetic challenge—selective C4 functionalization of pyridines, featuring operational\nsimplicity, transition metal-free conditions, and excellent air/moisture tolerance. The authors\nconvincingly demonstrate broad substrate scope, scalability, and mechanistic insight through\ncomplementary experimental and computational studies. By cleverly employing paired electrolysis,\nthe method achieves redox-neutral transformation with high step and atom economy. Overall, this\nis a well-executed and impactful study, I recommend its publication in Nature Communications after\nthe following minor points are addressed.\nAuthors’ Response: We sincerely appreciate your positive comments to our work.\n(1) Although TEMPO has been demonstrated to be an effective mediator in the oxidative\naromatization step, the authors should provide a brief explanation in the “Reaction Condition\nOptimization” section as to why alternative mediators, such as ferrocene, failed to facilitate the\ntransformation.\nResponse: Thank you for your suggestion. We have supplemented this section of the main text with\nan explanation clarifying why alternative mediators failed to facilitate this transformation. The\ndescriptions are as follows: “Neither ferrocene nor n-Bu NI enabled the formation of C1 (entries\n4\n10–11). This is attributed to the insufficient oxidizing capability of ferrocenium ions and elemental\niodine, which are unable to effectively promote the oxidation of the intermediate—a key step in\nproduct formation. Additionally, these mediators may undergo competitive side reactions, further\nreducing the overall reaction efficiency.”\n(2) The failure of cyclohexanone (B40) is noted, but no explanation is provided. Is this due to sterics,\nreduction potential, or radical stability? A brief comment would strengthen the scope discussion.\nResponse: Thank you for your suggestion. Cyclohexanone did not yield the desired product,\nprimarily because of its excessively high reduction potential. A brief comment to this effect has been\nincluded in the paragraph above Scheme 2 in the main text: “Notably, cyclohexanone (B40) did not\nreact productively; instead, the pyridine homocoupling product was obtained. This outcome stems\nfrom cyclohexanone's overly high reduction potential.”\n(3) For aldehydes, yields are generally moderate. Are there competing side reactions (e.g., over-\nreduction to alcohols or homo-coupling to vicinal diols)?\nResponse: Thank you for your suggestion. The low yield of aldehydes in the reaction is mainly due\nto their lower reduction potential compared with ketones, which leads to a mismatch with the\nreduction rate of pyridines substrates. As a result, over-reduction occurs in the reaction, producing\nbenzyl alcohols and small amount of vicinal diols.\n(4) The switch in regioselectivity for quinolines and phenanthroline (C2 vs. C4) is attributed to\nsterics/radical stability?\nResponse: Thank you for your insightful question. We observed C2-selectivity when using\nsubstrates such as 3-phenylpyridine, quinoline, and phenanthroline. In contrast, furo[3,2-b]pyridine\nand thieno[3,2-b]pyridine, which present less steric hindrance, afforded mixtures of C2- and C4-\nfunctionalized products (C57/C57' = 2.7:1; C58/C58' = 1.9:1). These experimental results indicate\nthat the difference in regioselectivity is mainly attributed to steric effects.\n(5) For the gram-scale reaction, were any adjustments (e.g., electrode size, stirring rate) necessary?\nResponse: Thank you for this meaningful question. The reaction at the 0.2 mmol scale was carried\nout under standard conditions using a 10 mL undivided electrolytic cell, with electrode plates\nmeasuring 5.25 cm × 0.8 cm × 0.2 cm and a stirring speed of 400 rpm. For the scaled-up reaction\n(3 mmol), a 30 mL undivided electrolytic cell was used, other parameters, including electrode plate\ndimensions, stirring speed, and inter-electrode distance, remained the same as in the standard setup.\n(6) The abbreviation “SCS” (spin center shift) should be defined upon first use.\nResponse: We have defined SCS, please see the paragraph above Scheme 2: “the latter suggests a\nspin center shift (SCS) of the hydroxy α-radical”.\n(7) In the introduction, references to “Scheme 1c1–1c3” should be checked for accuracy (currently\nlabeled as 1c1, 1c2, etc.).\nResponse: Thank you for pointing out this issue. In the introduction section of the main text, we\nhave revised it to avoid ambiguity.\n(8) The phrase “ubiquitous pyridine rings” could be rephrased to “ubiquitous pyridine scaffold” for\nprecision.\nResponse: Thank you for the suggestion. We have revised “ubiquitous pyridine rings” to\n“ubiquitous pyridine scaffold” in the abstract section.\n(9) Check formatting of references (e.g., reference 8e is missing, and 8f appears as 8g).\nResponse: Thank you for point out this error. We have corrected the formatting of all references to\nadhere strictly to Nature Communications guidelines.\nReviewer #2 (Remarks to the Author):\nThe manuscript describes a TEMPO-mediated paired electrolysis that delivers C4 hydroxyalkylated\npyridines from ketones and aldehydes at room temperature, with CV, control experiments, radical\nclocks, KIE, and DFT used to support a mechanism involving concurrent formation of pyridyl and\nhydroxy α radicals followed by oxidative aromatization. The overall concept—direct C4\nfunctionalization of pyridines with simple carbonyls under formally redox neutral conditions in an\nundivided cell—is attractive and addresses a relevant synthetic problem, although the extent of\nconceptual advance may be somewhat overstated.\nAuthors' Response: We sincerely thank you for your great effort in evaluation of our work and\nyour insightful comments.\n1.C4 selectivity and “inert pyridines”\nThe presentation of C4 selectivity and “inert pyridines” raises major concerns. Most of the high\nperforming examples are 2 aryl or otherwise 2 substituted pyridines, where both the DFT analysis\nand simple qualitative reasoning indicate that spin density is biased toward C4/C6; accordingly, the\nobserved C4 functionalization appears to be largely dictated by substrate electronics rather than by\na generally applicable site selectivity control element. The only example using unsubstituted\npyridine (C46) requires substantially modified conditions (graphite cathode, 35 mA, pyridine in\nexcess) and still gives only 40% yield with 10:1 C4:C2, suggesting that intrinsic C4 control on a\nsimple pyridine is modest and sensitive to conditions.\nSeveral substrates also give regioisomeric mixtures or even C2 preference (e.g., fused systems\nC54/C54′, C55/C55′), indicating that the method is not generally C4 selective across unbiased\npyridines. This contrasts with the title and abstract (“direct C4 hydroxyalkylation of inert pyridines”,\n“broad substrate scope”, “C4 selective”), which imply a more general and electronics agnostic\nplatform. It would therefore be helpful either to moderate these claims or to broaden the scope (e.g.,\nto simple 3 and 4 substituted and more diverse mono substituted pyridines) and provide full\nregioisomer profiles to support a claim of general C4 selectivity.\nResponse: Thank you very much for your insightful comments. Based on your suggestion, we have\nrevised the manuscript title to “Room-Temperature Direct Hydroxyalkylation of Pyridines via\nPaired Electrolysis,” removing the “C4-” designation.\n(1) We performed additional experiments using several representative pyridines, including\n4-(trifluoromethyl)pyridine (C ’), 3-(trifluoromethyl)pyridine (C ’), and 2-methylpyridine\n52 54\n(C ). The reaction is indeed influenced by the reduction potentials and steric effects of the\n47\nsubstrates, and its current scope is primarily centered on C2-substituted pyridines. As expected,\nwhen the C4 position is blocked, C4-substituted pyridines can react only at the C2 site (e.g.,\nproducts C ’ and C ’ in Fig.(cid:3031)1). Furthermore, steric hindrance directs C3-substituted pyridines\n51 52\ntoward C2-functionalization (see products C ’ and C ’). Consequently, the regioselectivity for\n53 54\nC3- and C4-substituted pyridines differs from that of unsubstituted (C ) and C2-substituted\n46\npyridines (see Fig. (cid:3031)2 and(cid:3031)Fig. 3 in the manuscript). The relatively low yield observed with\npyridines can be attributed to a mismatch in reduction potentials of two reactants.\n(2) Furthermore, the formation of regioisomers (e.g., fused systems C /C ′ and C /C ′) or\n54 54 55 55\ndistinct regioselectivity patterns observed with fused pyridine substrates can be attributed to\ntheir significantly different electronic and steric environments compared to standard\nmonocyclic pyridines. Similar trends have been noted in earlier studies on pyridine\nfunctionalization (J. Am. Chem. Soc. 2024, 146, 24257–24264; Nature 2023, 615, 67–72).\n(3) Although the regioselectivity of this reaction is partly influenced by substrate electronic effects,\nrecent studies on pyridine functionalization have also documented cases where regioselectivity\nis governed by substrate spin-density distributions (Nat. Synth. 2026, 5, 27–35). Notably, some\nof these reports demonstrate regioselectivity levels comparable to or even lower than those\nobserved in our work.\n2. Electrochemical concept: paired vs linear paired electrolysis\nThe electrochemical design does qualify as paired electrolysis in the usual synthetic sense: pyridine\nand the carbonyl compound are reduced at the cathode, while TEMPO is oxidized at the anode to\nTEMPO+, which then oxidizes the dihydropyridine intermediate to the aromatic product, with\nTEMPO/TEMPOH being regenerated so that both electrodes are used productively and no\nsacrificial anode or stoichiometric oxidant is required.\nHowever, the manuscript also uses the more specific term “linear paired electrolysis”, and this\nparticular designation is not clearly supported by the data. The discussion attributes “linear”\nbehavior to diffusion of the coupling adduct away from the cathodic double layer prior to anodic\noxidation, but the evidence for a dominant linear redox cycle remains qualitative: there is no\nquantitative demonstration that a single redox couple is continuously shuttling between electrodes\nor that the majority of current is carried by such a cycle (e.g., via faradaic efficiency, partitioning of\nanodic vs cathodic processes, potential profiles, or mediator mass balance). In this context,\ndescribing the system simply as “paired” or “redox neutral TEMPO mediated electrolysis” would\nseem more appropriate than emphasizing the stricter, mechanistically defined “linear paired” label.\nResponse: Thank you for your insightful comments. In response, we have revised the term “linear\npaired electrolysis” to “paired electrolysis” throughout the main text. Additionally, we have\ndetermined and calculated the corresponding Faradaic equivalents and Faradaic efficiency (see\nTable S1 in SI), as well as the reaction potential curve (Fig. S9), all of which are now provided in\nthe Supplementary Information.\n3. Missing quantitative electrochemical characterization Given how central “paired” and “redox\nneutral” electrolysis are to the narrative, the lack of faradaic efficiency and current efficiency data\nrepresents an important gap. It would be informative to see faradaic efficiencies for a few key\nexamples (ideally including the gram scale experiment) to support the redox neutral description and\nto indicate how much of the charge is directed into product formation rather than side processes\nsuch as overreduction, solvent/electrolyte oxidation, or unproductive mediator turnover, especially\nin the context of strong claims about efficient paired electrolysis.\nResponse: Thank you for your insightful comments. We have now supplemented the calculation of\nFaradaic efficiency for both the standard and the scaled-up reactions. The detailed calculation\nprocedures are provided as follows:\nstandard reaction:\nIt 25×10-3×9×3600\nn = = = 25.91 F/mol\nF znF 2×0.162×10-3×96485\n1 1\nFE = = = 3.86%\nn F 25.91\nscale-up reaction:\nIt 40×10-3×80×3600\nn = = = 32.09 F/mol\nF znF 2×1.86×10-3×96485\n1 1\nFE = = = 3.12%\nn F 32.09\nI: Reaction current (A); t: Reaction time (s); n: the moles of products (mol); F: Faraday’s constant\n(96485 C/mol). z: the number of electrons transferred per molecule. n : Faraday equivalent. FE:\nF\nFaraday efficiency.\nFollowing electrolysis under standard conditions, approximately 0.24 mmol of 4'-\nmethoxyacetophenone was recovered. Together with the aforementioned data, these results suggest\nthat the reaction is likely constrained by the relatively high reduction potential of the substrate and\nthe rate of radical cross-coupling. This limitation results in unproductive redox reactions of the\nsubstrate, as well as hydrogen and oxygen evolution side reactions in the undivided cell.\nAdditionally, preliminary attempts to employ alternating current electrolysis with the aim of\nenhancing the faradaic efficiency of the process were carried out, but no improvement was observed.\n4. Conceptual claims on “new reactivity paradigms”\nThe conclusion’s statement that merging reductive dearomatization with radical transformation\n“unlocks new reactivity paradigms for inert chemical systems” appears stronger than warranted by\nthe current data set. Related “dearomatize then functionalize/oxidize” strategies are already known,\nfor example in phosphonium/Zincke pyridinium chemistry, dihydropyridine hydride transfer, and\nother radical/polar dearomatization chemistry, and in this study the concept is mainly\ndemonstrated for electronically activated 2 substituted pyridines with limited exploration of unbiased\npyridines or other “inert” substrates. A more measured formulation (for example, indicating that the work\n“illustrates a strategy that may be extendable to other systems”) would better reflect what has been\nestablished experimentally.\nResponse: Thank you for your constructive comments and suggestions. While related strategies\ninvolving \"dearomatize-then-functionalize/oxidize\" sequences have been reported in earlier studies,\nthey typically involve stepwise operations. In contrast, the protocol described in this work proceeds\nin a single operation and requires no metal catalysts, stoichiometric redox reagents, or unstable\nagents. On this basis, we believe the present electrolysis-driven approach offers inherent advantages\nand distinct novelty. That said, we fully agree with your suggestion and have revised the relevant\nconclusion as follows: \"In this work, the strategy of performing electroreduction-induced\ndearomatization and subsequent intermediate transformation in a single operation is expected to\nadvance the development of efficient methodologies for (hetero)aryl functionalization.\")\n5. Supporting Information (NMR spectra)\nAs a minor point regarding the Supporting Information, the ¹H NMR spectra would benefit from\nexpanded views of the aromatic region; at present these regions are quite compressed, making it\ndifficult to clearly assess coupling patterns and independently verify the assigned substitution\npatterns.\nResponse: Thank you for your valuable feedbacks. As requested, enlarged versions of the product\n¹H NMR spectra have been provided. In these original spectra, signal overlap in the aromatic regions\nimpedes clear identification of coupling patterns.\n6. Overall assessment\nIn summary, the transformation is synthetically useful, and the general electrochemical strategy is\nof interest, but several central claims—general C4 selectivity across “inert” pyridines, the “linear\npaired electrolysis” designation, and the description of “new reactivity paradigms”—are not fully\nsupported by the current scope and data. The electrochemical characterization remains incomplete\n(no faradaic efficiencies and limited electroanalysis), and the substrate scope is heavily weighted\ntoward electronically biased 2 substituted pyridines, with only modest performance on truly\nunbiased pyridines. Taken together, these points suggest that, in its present form, the work does not\nyet reach the level of conceptual generality and mechanistic support expected for Nature\nCommunications, and may ultimately be better aligned with a more specialized journal once the\nclaims and presentation are adjusted accordingly.\nResponse: We sincerely thank you for your thoughtful and constructive evaluation of our work. We\nhave carefully considered all the points raised and have revised the manuscript accordingly.\n(1) Regarding regioselectivity and substrate scope: the majority of tested substrates exhibited\nC4/C2 selectivities above 10:1, demonstrating consistently high regiocontrol. This level of\nselectivity is in line with, or exceeds, that reported in recent leading studies on direct pyridine\nfunctionalization without directing groups (e.g., J. Am. Chem. Soc. 2024, 146, 24257–\n24264; Nat. Chem. 2021, 13, 1207–1213). We agree that extending the scope to fully unbiased\npyridines still remains challenging, largely due to inherent limitations imposed by substrate\nreduction potentials and less steric environment, which affect the efficiency of the radical cross-\ncoupling step. In response to this comment, we have expanded the substrate panel in the revised\nmanuscript and have more transparently discussed the factors influencing reactivity and\nselectivity.\n(2) On Terminology and Claims: Noteworthy, we have revised the manuscript title to “Room-\nTemperature Direct Hydroxyalkylation of Pyridines via Paired Electrolysis,” removing the “C4-”\ndesignation. Moreover, we have revised the term “linear paired electrolysis” throughout the text\nto the more accurate “paired electrolysis.” We have also moderated claims related to “new\nreactivity paradigms” and “general C4 selectivity,” reframing them to better reflect what the\nexperimental data support—namely, that this electrolysis-driven strategy offers a distinct and\npractical route for regioselective pyridine functionalization under transition metal catalyst -free\nconditions, and it overcomes the limitations of conventional pyridyl functionalization that\nrequire pre-functionalized substrates or stoichiometric activators.\n(3) Supplementary Electrochemical Data: As suggested, we have supplemented the Supporting\nInformation with detailed Faradaic efficiency calculations, reaction potential profiles, and\nadditional electrochemical characterization to provide a more complete analysis of the system.\n(4) Novelty and Contribution: while we recognize the need for clearer boundaries in our claims,\nwe believe the core advance—enabling direct, regioselective C–H cross-coupling of pyridines\nwith carbonyls in a single opernation, without metals, stoichiometric oxidants, or directing\ngroups—represents a meaningful and novel contribution to synthetic electrochemistry and\nheteroarene functionalization.\nWe are grateful for the opportunity to improve the manuscript and believe the revisions have\nstrengthened its clarity, accuracy, and scholarly impact. We hope the revised version now meets the\nhigh standards expected by the journal.\nReviewer #3 (Remarks to the Author):\nIn this manuscript, Zhang and co-workers report an electroreductive C–C bond formation between\npyridine derivatives and carbonyl compounds. By leveraging paired electrolysis, the authors achieve\ndirect C4-functionalization of pyridines under mild acidic conditions without the need for pre-\nactivation. This transformation employs operationally practical, air- and moisture-tolerant\nconditions, representing an innovative and impactful example of electrochemical radical–radical\ncoupling. The manuscript is well-structured and clearly written. The electrochemical protocols are\ndescribed in detail and have been successfully applied to the late-stage functionalization of complex\nbioactive molecules, underscoring the practical utility of the method. Given the novelty,\nmethodological rigor, and scientific significance of this work, I recommend the manuscript for\npublication in Nature Communications following the minor revisions listed below.\nAuthors' Response: We sincerely thank you for your encouraging and insightful comments on our\nwork.\n1) What exactly happened when acetone was used as the solvent, were any byproducts observed?\nResponse: Thank you for your interesting question. When acetone was used as the solvent, a\nsignificant portion of pyridine starting material remained unreacted, leading to a mixture of C4- and\nC2-functionalized products in an approximate ratio of 3:1 with an overall low yield. Additionally, a\nsmall amount of vicinal diol by-product was detected.\n2) In this work, the reaction is achieved by employing TEMPO as a redox mediator. Can the reaction\nbe achieved in high yield through a simple workup after completing the electrolysis with the\ncondition employing a zinc anode?\nResponse: Thank you for raising this interesting point. When a zinc anode was used, the target\nproduct was obtained in 15% yield. However, a significant portion of the pyridine starting material\nremained unreacted, whereas 4′-methoxyacetophenone was nearly fully consumed and converted\ninto pinacol by-product. Attempts to improve the product yield through post-treatment methods,\nsuch as adding oxidants, were unsuccessful.\n3) Reaction times are reported in the optimization of reaction conditions. However, faradaic\nequivalents (F/mol) provide a more informative measure of efficiency than reaction time, enabling\na marked improvement in the reaction's reproducibility.\nResponse: Thank you for your helpful suggestion. In response, we have calculated the Faradaic\nequivalent and Faradaic efficiency for the model reaction and included the results in the Supporting\nInformation (Table S1). The detailed calculation procedure is provided below.\nI: Reaction current (A); t: Reaction time (s); n: the moles of products (mol); F: Faraday’s constant\n(96485 C/mol). z: the number of electrons transferred per molecule. n : Faraday equivalent. FE:\nF\nFaraday efficiency.\n4) Was the excess acetophenone fully consumed and converted into by-products during the reaction?\nResponse: Thank you for your question. During the reaction, some acetophenone was converted\ninto pinacol by-products, while some remained unreacted.\n5) In terms of the mechanism, it is very challenging to have two different radicals to combine due\nto the low concentration and short lifetime. The DFT calculation further illustrates the feasibility of\nthe mechanism. Can the authors explain why the alkyl ketones do not proceed smoothly?\nResponse: Thank you for raising this important question. The α-alkyl radicals generated from\nelectron-deficient alkyl ketones gave the desired product in moderate yield (see compound C22). In\ncontrast, electron-rich alkyl ketones failed to yield any target products, primarily due to their\nexcessively high reduction potentials, which lead to a kinetic mismatch between the reduction and\ncoupling steps.\nReviewer #4 (Remarks to the Author):\nIn this work, Zhang and coworkers developed an innovative acetic acid-activated electrolysis\nstrategy for direct C4-functionalization of inert pyridines with ketones/imines at room temperature.\nThis method enables the introduction of electrophiles into pyridine scaffolds without pre-\nfunctionalization steps, demonstrating good functional group tolerance and broad synthetic utility.\nMechanistic investigations, including experimental studies and DFT calculations, provide insights\ninto the reaction mechanism and chemoselectivity. Overall, this work presents an efficient\nelectrochemical approach for C4-functionalization of inert pyridines; however, several issues\nrequire clarification before further consideration.\nAuthors’ Response: We sincerely appreciate your positive feedback and constructive suggestions\non our manuscript.\n1. Table 1 shows TEMPO as a reaction component, while Figure 1A-III indicates TEMPO inhibits\nproduct formation. This apparent contradiction requires explanation.\nResponse: Thank you for your question regarding the role of TEMPO in this reaction. To clarify,\nTEMPO acts as a mediator to promote aromatization through oxidation. However, at higher\nconcentrations, it functions as a radical quencher, thereby suppressing the formation of the desired\nproduct. This concentration-dependent dual role of TEMPO has been documented in prior studies\n(Green Chem., 2022, 24, 4928-4934).\n2. In Figure 1A, the authors mentioned that homocoupling occurs when A1 or B1 is reacted\nindividually. However, under standard conditions where both substrates are present simultaneously,\nheterocoupling is exclusively favored with no detectable homocoupling products. This selectivity\npattern requires mechanistic elucidation. The authors are encouraged to perform computational\nstudies to provide a reasonable explanation.\nResponse: Thank you for your suggestions. We would like to clarify that under the standard reaction\nconditions, the system does generate homocoupling products; in fact, a small amount of pinacol\nby-product is also formed during the reaction. Furthermore, when 3 equivalents of 2-phenylpyridine\nand 1 equivalent of 4-methoxyacetophenone were used (Table S1, entry(cid:3031)20), the target product was\nobtained in 57% yield, accompanied by the pyridine homocoupling product in approximately 17%\nyield (yield calculated based on pyridine equivalents). The cross-coupling mechanism in this\nreaction depends mainly on the electronic compatibility between the two distinct radicals, and is\nalso influenced by the rate of radical generation—consistent with our previous reports\n(J.(cid:3031)Am.(cid:3031)Chem.(cid:3031)Soc.(cid:3031)2023,(cid:3031)145,(cid:3031)10967–10973; Org.(cid:3031)Lett.(cid:3031)2025,(cid:3031)27,(cid:3031)4725–4731).\n3. The KIE in Figure 1C is interpreted as evidence for rate-determining C–H cleavage. However,\nfollowing established literature (Angew. Chem. Int. Ed. 2012, 51, 3066), KIE data alone may not\ndefinitively identify the RDS. The text should be revised to acknowledge alternative interpretations.\nResponse: Thank you for your comments. We have revised the expression as follows:“This result\nsuggests that C–H bond cleavage at the pyridyl C4 position is likely involved in the rate-determining\nstep, though the presence of other rate-limiting processes cannot be excluded.” The reference is\ncited as ref 77.\n4. Please clarify: (1) Is \"M06-2x-dd3\" in Figure 2 a typo? (2) Why were different solvation models\n(PCM for optimization, SMD for energies) employed? Reference support for this approach should\nbe provided.\nResponse: (1) Yes. “M06-2x-dd3” in Figure 2 is a typographical error. It has been corrected to M06-\n2X-D3 in the revised figure and associated text.\n(2) We used IEFPCM (PCM) for geometry optimizations and frequency analyses mainly for\nnumerical robustness. Employing SMD during optimization/frequency calculations can\noccasionally make convergence more difficult and may introduce small symmetry-breaking\ndistortions (especially for high-symmetry stationary points), which can lead to spurious\nlow/imaginary frequencies. By contrast, IEFPCM is generally more stable for these tasks, and\nGaussian has specifically improved PCM optimization behavior in recent versions. It is also\ncommonly to use PCM solvent model for optimization and SMD solvent model for high level single\npoint calculation in literatures. (see, Nat. Commun. 16, 10283, (2025); Angew. Chem. Int. Ed. 64,\ne202501924, (2025))\n5. The authors claim “the electron spin predominantly located on the C4 and C6 positions, with little\nspin density at the C2 position.” However, only the spin populations at C2 and C4 are labeled in\nFigure 1. It is suggested to label the spin populations at N and C6 as well.\nResponse: Thank you for the suggestions. We agree that explicitly labeling the spin populations at\nN and C6 would make the statement clearer and improve the readability of figure. In the revised\nmanuscript, we have added the spin population labels at N and C6 and updated the figure\naccordingly.\n)lom/lack(\nygrenE\nevitaleR\nFigure 3\n6. Figure S10D is identical to Figure 1. Please remove this redundant figure.\nResponse: Thank you for your suggestion. The redundant figure has now been removed."}, {"round": 2, "reviewer_comments": {"reviewer_1": "The author has addressed all the concerns raised by the reviewers.Therefore, I recommend acceptance of the manuscript in its present form.", "reviewer_2": "I appreciate the authors' efforts in addressing the previous concerns. They have added new substrates, provided Faradaic efficiency data, and made adjustments to the title and terminology. However, several important issues remain that need clarification before the manuscript meets the standards expected for Nature Communications.\nThe work presents a useful electrochemical method for functionalizing pyridines under mild, metal-free conditions. The method works best for 2-aryl substituted pyridines and can be extended to simpler pyridines and other heterocycles, though with lower yields and harsher conditions. The mechanistic studies are informative, and the practical applications are valuable. My concerns are primarily about presentation and ensuring claims align with the actual data.\nThe following points require attention:\n1. Mismatch Between Title and Narrative\nWhile the title was changed to remove \"C4-\", the introduction and main narrative still heavily emphasize C4-functionalization of \"inert pyridines.\" This creates a fundamental disconnect: the title suggests a general method for pyridine hydroxyalkylation, but the text continues to emphasize C4-selectivity that is actually substrate-dependent rather than method-controlled.\nAdditionally, the term \"paired electrolysis\" in the title may overstate the efficiency of the system given the very low Faradaic efficiency (3.86%, see point 4 below), which suggests that paired electrolysis is not the dominant pathway.\nSuggestion: The title and narrative need to be aligned to accurately represent the scope and nature of the work. Options include: (a) revise the introduction to match the current title, clearly stating that the method works best for 2-substituted pyridines where substrate electronics favor C4-selectivity, and discuss the low FE honestly; OR (b) change the title to more accurately reflect the actual scope and mechanism, such as \"Room-Temperature Electrochemical C4-Hydroxyalkylation of 2-Arylpyridines\" (removing \"paired electrolysis\" claim and specifying substrate class). The key issue is consistency—the title, narrative, and data should all align to give readers an accurate understanding of what has been achieved.\n2. Scope and \"Inert Pyridines\" Claims\nThe newly added substrates show the limitations more clearly:\n• C46 (pyridine): 40% yield with modified conditions\n• C47 (2-methylpyridine): 24% yield\n• C52' (4-CF₃-pyridine): 29% yield, C2-selective\n• C54' (3-CF₃-pyridine): 31% yield, C2-selective\nThese results indicate the method is optimized for 2-aryl substituted pyridines rather than being generally applicable to \"inert pyridines.\" The DFT calculations support this: C4-selectivity arises primarily from substrate electronics (the 2-aryl substituent) combined with steric effects, not from the electrochemical method itself. This is substrate-controlled selectivity, not method-controlled selectivity.\nSuggestion: Please clarify in the text that the method works best for 2-aryl (or 2-heteroaryl) substituted pyridines. The \"inert pyridines\" framing should be softened to accurately represent the scope and avoid overstating the generality.\n3. Comparative Claims Need Support\nIn the response letter, you state that selectivity \"is in line with, or exceeds\" recent studies (citing JACS 2024, 146, 24257 and Nat. Chem. 2021, 13, 1207). However, this comparison does not appear in the manuscript. If your regioselectivity is indeed improved compared to these methods, showing this comparison would strengthen your work. Additionally, explaining why your method achieves better regioselectivity (if applicable) would be valuable.\nSuggestion: Either add a brief comparative table or discussion showing how your method compares to these references, or remove the comparative claim. If you choose to include the comparison, please explain the basis for any superiority in regioselectivity.\n4. Faradaic Efficiency Discussion Missing\nThe calculated Faradaic efficiencies (3.86% standard, 3.12% scale-up) are quite low, indicating that ~96% of electrons are consumed by side reactions rather than product formation. This raises significant concerns about the \"high efficiency\" and \"paired electrolysis\" claims. For context, well-functioning paired electrolysis systems typically show FE >30%.\nI understand that low FE can occur in undivided cells with competing H₂/O₂ evolution, but this should be discussed transparently in the main text rather than just reported in the SI. The low FE directly contradicts the \"efficient paired electrolysis\" narrative presented throughout the manuscript.\nSuggestion: Add a paragraph discussing the FE values and their implications for the efficiency claims. Either explain why low FE is acceptable for this system (e.g., operational simplicity outweighs electron efficiency), or acknowledge it as a limitation that could be improved in future work. Claims about \"high atom economy\" and \"efficient paired electrolysis\" should be qualified or removed given the 3.86% FE.\n5. NMR Spectra Expansion Incomplete\nWhile some NMR spectra now show expanded aromatic regions, many still remain compressed. Please provide systematic expansion of aromatic regions for all products to allow verification of structural assignments.\nOverall Assessment\nThis work demonstrates several valuable features:\n• A metal-free electrochemical approach for pyridine functionalization\n• Room temperature, operationally simple conditions\n• Mechanistic insights through CV, DFT, and control experiments\n• Practical utility for late-stage functionalization of 2-aryl pyridines\n• Air and moisture tolerance\nThese are legitimate contributions. However, the presentation needs substantial adjustment to ensure claims accurately reflect the data:\n• Scope: More specific than \"inert pyridines\"—works best for 2-aryl substituted pyridines, with poorer performance for simpler pyridines\n• Efficiency: Lower than suggested by current claims (3.86% FE needs honest discussion)\n• Regioselectivity: Substrate-dependent (electronic and steric effects), not method-controlled\n• Generality: The \"inert pyridine\" framing oversells the actual scope\nNature Communications readers expect rigorous, honest reporting. With appropriate revisions addressing the five points above, this work should be presented as \"an electrochemical method for hydroxyalkylation of 2-aryl substituted pyridines under mild, metal-free conditions\" rather than as a general solution for functionalizing \"inert pyridines\" with \"high efficiency.\"\nIf the authors can address these presentation issues and align their claims with the actual data, the manuscript could make a useful contribution to the electrochemical synthesis literature.", "reviewer_3": "Previous comments have been properly addressed. This work is recommended for publication with Nature Communications.", "reviewer_4": "The authors have addressed the questions reasonably well and I recommend the manuscript for publication in Nature Communications."}, "author_response": "Point-by-point response to Reviewer comments\nREVIEWER COMMENTS\nReviewer #1 (Remarks to the Author):\nThis manuscript presents a novel and elegant electrochemical strategy for the direct C4-\nhydroxyalkylation of pyridines using readily available carbonyl compounds. The work addresses a\nsignificant synthetic challenge—selective C4 functionalization of pyridines, featuring operational\nsimplicity, transition metal-free conditions, and excellent air/moisture tolerance. The authors\nconvincingly demonstrate broad substrate scope, scalability, and mechanistic insight through\ncomplementary experimental and computational studies. By cleverly employing paired electrolysis,\nthe method achieves redox-neutral transformation with high step and atom economy. Overall, this\nis a well-executed and impactful study, I recommend its publication in Nature Communications after\nthe following minor points are addressed.\nAuthors’ Response: We sincerely appreciate your positive comments to our work.\n(1) Although TEMPO has been demonstrated to be an effective mediator in the oxidative\naromatization step, the authors should provide a brief explanation in the “Reaction Condition\nOptimization” section as to why alternative mediators, such as ferrocene, failed to facilitate the\ntransformation.\nResponse: Thank you for your suggestion. We have supplemented this section of the main text with\nan explanation clarifying why alternative mediators failed to facilitate this transformation. The\ndescriptions are as follows: “Neither ferrocene nor n-Bu NI enabled the formation of C1 (entries\n4\n10–11). This is attributed to the insufficient oxidizing capability of ferrocenium ions and elemental\niodine, which are unable to effectively promote the oxidation of the intermediate—a key step in\nproduct formation. Additionally, these mediators may undergo competitive side reactions, further\nreducing the overall reaction efficiency.”\n(2) The failure of cyclohexanone (B40) is noted, but no explanation is provided. Is this due to sterics,\nreduction potential, or radical stability? A brief comment would strengthen the scope discussion.\nResponse: Thank you for your suggestion. Cyclohexanone did not yield the desired product,\nprimarily because of its excessively high reduction potential. A brief comment to this effect has been\nincluded in the paragraph above Scheme 2 in the main text: “Notably, cyclohexanone (B40) did not\nreact productively; instead, the pyridine homocoupling product was obtained. This outcome stems\nfrom cyclohexanone's overly high reduction potential.”\n(3) For aldehydes, yields are generally moderate. Are there competing side reactions (e.g., over-\nreduction to alcohols or homo-coupling to vicinal diols)?\nResponse: Thank you for your suggestion. The low yield of aldehydes in the reaction is mainly due\nto their lower reduction potential compared with ketones, which leads to a mismatch with the\nreduction rate of pyridines substrates. As a result, over-reduction occurs in the reaction, producing\nbenzyl alcohols and small amount of vicinal diols.\n(4) The switch in regioselectivity for quinolines and phenanthroline (C2 vs. C4) is attributed to\nsterics/radical stability?\nResponse: Thank you for your insightful question. We observed C2-selectivity when using\nsubstrates such as 3-phenylpyridine, quinoline, and phenanthroline. In contrast, furo[3,2-b]pyridine\nand thieno[3,2-b]pyridine, which present less steric hindrance, afforded mixtures of C2- and C4-\nfunctionalized products (C57/C57' = 2.7:1; C58/C58' = 1.9:1). These experimental results indicate\nthat the difference in regioselectivity is mainly attributed to steric effects.\n(5) For the gram-scale reaction, were any adjustments (e.g., electrode size, stirring rate) necessary?\nResponse: Thank you for this meaningful question. The reaction at the 0.2 mmol scale was carried\nout under standard conditions using a 10 mL undivided electrolytic cell, with electrode plates\nmeasuring 5.25 cm × 0.8 cm × 0.2 cm and a stirring speed of 400 rpm. For the scaled-up reaction\n(3 mmol), a 30 mL undivided electrolytic cell was used, other parameters, including electrode plate\ndimensions, stirring speed, and inter-electrode distance, remained the same as in the standard setup.\n(6) The abbreviation “SCS” (spin center shift) should be defined upon first use.\nResponse: We have defined SCS, please see the paragraph above Scheme 2: “the latter suggests a\nspin center shift (SCS) of the hydroxy α-radical”.\n(7) In the introduction, references to “Scheme 1c1–1c3” should be checked for accuracy (currently\nlabeled as 1c1, 1c2, etc.).\nResponse: Thank you for pointing out this issue. In the introduction section of the main text, we\nhave revised it to avoid ambiguity.\n(8) The phrase “ubiquitous pyridine rings” could be rephrased to “ubiquitous pyridine scaffold” for\nprecision.\nResponse: Thank you for the suggestion. We have revised “ubiquitous pyridine rings” to\n“ubiquitous pyridine scaffold” in the abstract section.\n(9) Check formatting of references (e.g., reference 8e is missing, and 8f appears as 8g).\nResponse: Thank you for point out this error. We have corrected the formatting of all references to\nadhere strictly to Nature Communications guidelines.\nReviewer #2 (Remarks to the Author):\nThe manuscript describes a TEMPO-mediated paired electrolysis that delivers C4 hydroxyalkylated\npyridines from ketones and aldehydes at room temperature, with CV, control experiments, radical\nclocks, KIE, and DFT used to support a mechanism involving concurrent formation of pyridyl and\nhydroxy α radicals followed by oxidative aromatization. The overall concept—direct C4\nfunctionalization of pyridines with simple carbonyls under formally redox neutral conditions in an\nundivided cell—is attractive and addresses a relevant synthetic problem, although the extent of\nconceptual advance may be somewhat overstated.\nAuthors' Response: We sincerely thank you for your great effort in evaluation of our work and\nyour insightful comments.\n1.C4 selectivity and “inert pyridines”\nThe presentation of C4 selectivity and “inert pyridines” raises major concerns. Most of the high\nperforming examples are 2 aryl or otherwise 2 substituted pyridines, where both the DFT analysis\nand simple qualitative reasoning indicate that spin density is biased toward C4/C6; accordingly, the\nobserved C4 functionalization appears to be largely dictated by substrate electronics rather than by\na generally applicable site selectivity control element. The only example using unsubstituted\npyridine (C46) requires substantially modified conditions (graphite cathode, 35 mA, pyridine in\nexcess) and still gives only 40% yield with 10:1 C4:C2, suggesting that intrinsic C4 control on a\nsimple pyridine is modest and sensitive to conditions.\nSeveral substrates also give regioisomeric mixtures or even C2 preference (e.g., fused systems\nC54/C54′, C55/C55′), indicating that the method is not generally C4 selective across unbiased\npyridines. This contrasts with the title and abstract (“direct C4 hydroxyalkylation of inert pyridines”,\n“broad substrate scope”, “C4 selective”), which imply a more general and electronics agnostic\nplatform. It would therefore be helpful either to moderate these claims or to broaden the scope (e.g.,\nto simple 3 and 4 substituted and more diverse mono substituted pyridines) and provide full\nregioisomer profiles to support a claim of general C4 selectivity.\nResponse: Thank you very much for your insightful comments. Based on your suggestion, we have\nrevised the manuscript title to “Room-Temperature Direct Hydroxyalkylation of Pyridines via\nPaired Electrolysis,” removing the “C4-” designation.\n(1) We performed additional experiments using several representative pyridines, including\n4-(trifluoromethyl)pyridine (C ’), 3-(trifluoromethyl)pyridine (C ’), and 2-methylpyridine\n52 54\n(C ). The reaction is indeed influenced by the reduction potentials and steric effects of the\n47\nsubstrates, and its current scope is primarily centered on C2-substituted pyridines. As expected,\nwhen the C4 position is blocked, C4-substituted pyridines can react only at the C2 site (e.g.,\nproducts C ’ and C ’ in Fig.(cid:3031)1). Furthermore, steric hindrance directs C3-substituted pyridines\n51 52\ntoward C2-functionalization (see products C ’ and C ’). Consequently, the regioselectivity for\n53 54\nC3- and C4-substituted pyridines differs from that of unsubstituted (C ) and C2-substituted\n46\npyridines (see Fig. (cid:3031)2 and(cid:3031)Fig. 3 in the manuscript). The relatively low yield observed with\npyridines can be attributed to a mismatch in reduction potentials of two reactants.\n(2) Furthermore, the formation of regioisomers (e.g., fused systems C /C ′ and C /C ′) or\n54 54 55 55\ndistinct regioselectivity patterns observed with fused pyridine substrates can be attributed to\ntheir significantly different electronic and steric environments compared to standard\nmonocyclic pyridines. Similar trends have been noted in earlier studies on pyridine\nfunctionalization (J. Am. Chem. Soc. 2024, 146, 24257–24264; Nature 2023, 615, 67–72).\n(3) Although the regioselectivity of this reaction is partly influenced by substrate electronic effects,\nrecent studies on pyridine functionalization have also documented cases where regioselectivity\nis governed by substrate spin-density distributions (Nat. Synth. 2026, 5, 27–35). Notably, some\nof these reports demonstrate regioselectivity levels comparable to or even lower than those\nobserved in our work.\n2. Electrochemical concept: paired vs linear paired electrolysis\nThe electrochemical design does qualify as paired electrolysis in the usual synthetic sense: pyridine\nand the carbonyl compound are reduced at the cathode, while TEMPO is oxidized at the anode to\nTEMPO+, which then oxidizes the dihydropyridine intermediate to the aromatic product, with\nTEMPO/TEMPOH being regenerated so that both electrodes are used productively and no\nsacrificial anode or stoichiometric oxidant is required.\nHowever, the manuscript also uses the more specific term “linear paired electrolysis”, and this\nparticular designation is not clearly supported by the data. The discussion attributes “linear”\nbehavior to diffusion of the coupling adduct away from the cathodic double layer prior to anodic\noxidation, but the evidence for a dominant linear redox cycle remains qualitative: there is no\nquantitative demonstration that a single redox couple is continuously shuttling between electrodes\nor that the majority of current is carried by such a cycle (e.g., via faradaic efficiency, partitioning of\nanodic vs cathodic processes, potential profiles, or mediator mass balance). In this context,\ndescribing the system simply as “paired” or “redox neutral TEMPO mediated electrolysis” would\nseem more appropriate than emphasizing the stricter, mechanistically defined “linear paired” label.\nResponse: Thank you for your insightful comments. In response, we have revised the term “linear\npaired electrolysis” to “paired electrolysis” throughout the main text. Additionally, we have\ndetermined and calculated the corresponding Faradaic equivalents and Faradaic efficiency (see\nTable S1 in SI), as well as the reaction potential curve (Fig. S9), all of which are now provided in\nthe Supplementary Information.\n3. Missing quantitative electrochemical characterization Given how central “paired” and “redox\nneutral” electrolysis are to the narrative, the lack of faradaic efficiency and current efficiency data\nrepresents an important gap. It would be informative to see faradaic efficiencies for a few key\nexamples (ideally including the gram scale experiment) to support the redox neutral description and\nto indicate how much of the charge is directed into product formation rather than side processes\nsuch as overreduction, solvent/electrolyte oxidation, or unproductive mediator turnover, especially\nin the context of strong claims about efficient paired electrolysis.\nResponse: Thank you for your insightful comments. We have now supplemented the calculation of\nFaradaic efficiency for both the standard and the scaled-up reactions. The detailed calculation\nprocedures are provided as follows:\nstandard reaction:\nIt 25×10-3×9×3600\nn = = = 25.91 F/mol\nF znF 2×0.162×10-3×96485\n1 1\nFE = = = 3.86%\nn F 25.91\nscale-up reaction:\nIt 40×10-3×80×3600\nn = = = 32.09 F/mol\nF znF 2×1.86×10-3×96485\n1 1\nFE = = = 3.12%\nn F 32.09\nI: Reaction current (A); t: Reaction time (s); n: the moles of products (mol); F: Faraday’s constant\n(96485 C/mol). z: the number of electrons transferred per molecule. n : Faraday equivalent. FE:\nF\nFaraday efficiency.\nFollowing electrolysis under standard conditions, approximately 0.24 mmol of 4'-\nmethoxyacetophenone was recovered. Together with the aforementioned data, these results suggest\nthat the reaction is likely constrained by the relatively high reduction potential of the substrate and\nthe rate of radical cross-coupling. This limitation results in unproductive redox reactions of the\nsubstrate, as well as hydrogen and oxygen evolution side reactions in the undivided cell.\nAdditionally, preliminary attempts to employ alternating current electrolysis with the aim of\nenhancing the faradaic efficiency of the process were carried out, but no improvement was observed.\n4. Conceptual claims on “new reactivity paradigms”\nThe conclusion’s statement that merging reductive dearomatization with radical transformation\n“unlocks new reactivity paradigms for inert chemical systems” appears stronger than warranted by\nthe current data set. Related “dearomatize then functionalize/oxidize” strategies are already known,\nfor example in phosphonium/Zincke pyridinium chemistry, dihydropyridine hydride transfer, and\nother radical/polar dearomatization chemistry, and in this study the concept is mainly\ndemonstrated for electronically activated 2 substituted pyridines with limited exploration of unbiased\npyridines or other “inert” substrates. A more measured formulation (for example, indicating that the work\n“illustrates a strategy that may be extendable to other systems”) would better reflect what has been\nestablished experimentally.\nResponse: Thank you for your constructive comments and suggestions. While related strategies\ninvolving \"dearomatize-then-functionalize/oxidize\" sequences have been reported in earlier studies,\nthey typically involve stepwise operations. In contrast, the protocol described in this work proceeds\nin a single operation and requires no metal catalysts, stoichiometric redox reagents, or unstable\nagents. On this basis, we believe the present electrolysis-driven approach offers inherent advantages\nand distinct novelty. That said, we fully agree with your suggestion and have revised the relevant\nconclusion as follows: \"In this work, the strategy of performing electroreduction-induced\ndearomatization and subsequent intermediate transformation in a single operation is expected to\nadvance the development of efficient methodologies for (hetero)aryl functionalization.\")\n5. Supporting Information (NMR spectra)\nAs a minor point regarding the Supporting Information, the ¹H NMR spectra would benefit from\nexpanded views of the aromatic region; at present these regions are quite compressed, making it\ndifficult to clearly assess coupling patterns and independently verify the assigned substitution\npatterns.\nResponse: Thank you for your valuable feedbacks. As requested, enlarged versions of the product\n¹H NMR spectra have been provided. In these original spectra, signal overlap in the aromatic regions\nimpedes clear identification of coupling patterns.\n6. Overall assessment\nIn summary, the transformation is synthetically useful, and the general electrochemical strategy is\nof interest, but several central claims—general C4 selectivity across “inert” pyridines, the “linear\npaired electrolysis” designation, and the description of “new reactivity paradigms”—are not fully\nsupported by the current scope and data. The electrochemical characterization remains incomplete\n(no faradaic efficiencies and limited electroanalysis), and the substrate scope is heavily weighted\ntoward electronically biased 2 substituted pyridines, with only modest performance on truly\nunbiased pyridines. Taken together, these points suggest that, in its present form, the work does not\nyet reach the level of conceptual generality and mechanistic support expected for Nature\nCommunications, and may ultimately be better aligned with a more specialized journal once the\nclaims and presentation are adjusted accordingly.\nResponse: We sincerely thank you for your thoughtful and constructive evaluation of our work. We\nhave carefully considered all the points raised and have revised the manuscript accordingly.\n(1) Regarding regioselectivity and substrate scope: the majority of tested substrates exhibited\nC4/C2 selectivities above 10:1, demonstrating consistently high regiocontrol. This level of\nselectivity is in line with, or exceeds, that reported in recent leading studies on direct pyridine\nfunctionalization without directing groups (e.g., J. Am. Chem. Soc. 2024, 146, 24257–\n24264; Nat. Chem. 2021, 13, 1207–1213). We agree that extending the scope to fully unbiased\npyridines still remains challenging, largely due to inherent limitations imposed by substrate\nreduction potentials and less steric environment, which affect the efficiency of the radical cross-\ncoupling step. In response to this comment, we have expanded the substrate panel in the revised\nmanuscript and have more transparently discussed the factors influencing reactivity and\nselectivity.\n(2) On Terminology and Claims: Noteworthy, we have revised the manuscript title to “Room-\nTemperature Direct Hydroxyalkylation of Pyridines via Paired Electrolysis,” removing the “C4-”\ndesignation. Moreover, we have revised the term “linear paired electrolysis” throughout the text\nto the more accurate “paired electrolysis.” We have also moderated claims related to “new\nreactivity paradigms” and “general C4 selectivity,” reframing them to better reflect what the\nexperimental data support—namely, that this electrolysis-driven strategy offers a distinct and\npractical route for regioselective pyridine functionalization under transition metal catalyst -free\nconditions, and it overcomes the limitations of conventional pyridyl functionalization that\nrequire pre-functionalized substrates or stoichiometric activators.\n(3) Supplementary Electrochemical Data: As suggested, we have supplemented the Supporting\nInformation with detailed Faradaic efficiency calculations, reaction potential profiles, and\nadditional electrochemical characterization to provide a more complete analysis of the system.\n(4) Novelty and Contribution: while we recognize the need for clearer boundaries in our claims,\nwe believe the core advance—enabling direct, regioselective C–H cross-coupling of pyridines\nwith carbonyls in a single opernation, without metals, stoichiometric oxidants, or directing\ngroups—represents a meaningful and novel contribution to synthetic electrochemistry and\nheteroarene functionalization.\nWe are grateful for the opportunity to improve the manuscript and believe the revisions have\nstrengthened its clarity, accuracy, and scholarly impact. We hope the revised version now meets the\nhigh standards expected by the journal.\nReviewer #3 (Remarks to the Author):\nIn this manuscript, Zhang and co-workers report an electroreductive C–C bond formation between\npyridine derivatives and carbonyl compounds. By leveraging paired electrolysis, the authors achieve\ndirect C4-functionalization of pyridines under mild acidic conditions without the need for pre-\nactivation. This transformation employs operationally practical, air- and moisture-tolerant\nconditions, representing an innovative and impactful example of electrochemical radical–radical\ncoupling. The manuscript is well-structured and clearly written. The electrochemical protocols are\ndescribed in detail and have been successfully applied to the late-stage functionalization of complex\nbioactive molecules, underscoring the practical utility of the method. Given the novelty,\nmethodological rigor, and scientific significance of this work, I recommend the manuscript for\npublication in Nature Communications following the minor revisions listed below.\nAuthors' Response: We sincerely thank you for your encouraging and insightful comments on our\nwork.\n1) What exactly happened when acetone was used as the solvent, were any byproducts observed?\nResponse: Thank you for your interesting question. When acetone was used as the solvent, a\nsignificant portion of pyridine starting material remained unreacted, leading to a mixture of C4- and\nC2-functionalized products in an approximate ratio of 3:1 with an overall low yield. Additionally, a\nsmall amount of vicinal diol by-product was detected.\n2) In this work, the reaction is achieved by employing TEMPO as a redox mediator. Can the reaction\nbe achieved in high yield through a simple workup after completing the electrolysis with the\ncondition employing a zinc anode?\nResponse: Thank you for raising this interesting point. When a zinc anode was used, the target\nproduct was obtained in 15% yield. However, a significant portion of the pyridine starting material\nremained unreacted, whereas 4′-methoxyacetophenone was nearly fully consumed and converted\ninto pinacol by-product. Attempts to improve the product yield through post-treatment methods,\nsuch as adding oxidants, were unsuccessful.\n3) Reaction times are reported in the optimization of reaction conditions. However, faradaic\nequivalents (F/mol) provide a more informative measure of efficiency than reaction time, enabling\na marked improvement in the reaction's reproducibility.\nResponse: Thank you for your helpful suggestion. In response, we have calculated the Faradaic\nequivalent and Faradaic efficiency for the model reaction and included the results in the Supporting\nInformation (Table S1). The detailed calculation procedure is provided below.\nI: Reaction current (A); t: Reaction time (s); n: the moles of products (mol); F: Faraday’s constant\n(96485 C/mol). z: the number of electrons transferred per molecule. n : Faraday equivalent. FE:\nF\nFaraday efficiency.\n4) Was the excess acetophenone fully consumed and converted into by-products during the reaction?\nResponse: Thank you for your question. During the reaction, some acetophenone was converted\ninto pinacol by-products, while some remained unreacted.\n5) In terms of the mechanism, it is very challenging to have two different radicals to combine due\nto the low concentration and short lifetime. The DFT calculation further illustrates the feasibility of\nthe mechanism. Can the authors explain why the alkyl ketones do not proceed smoothly?\nResponse: Thank you for raising this important question. The α-alkyl radicals generated from\nelectron-deficient alkyl ketones gave the desired product in moderate yield (see compound C22). In\ncontrast, electron-rich alkyl ketones failed to yield any target products, primarily due to their\nexcessively high reduction potentials, which lead to a kinetic mismatch between the reduction and\ncoupling steps.\nReviewer #4 (Remarks to the Author):\nIn this work, Zhang and coworkers developed an innovative acetic acid-activated electrolysis\nstrategy for direct C4-functionalization of inert pyridines with ketones/imines at room temperature.\nThis method enables the introduction of electrophiles into pyridine scaffolds without pre-\nfunctionalization steps, demonstrating good functional group tolerance and broad synthetic utility.\nMechanistic investigations, including experimental studies and DFT calculations, provide insights\ninto the reaction mechanism and chemoselectivity. Overall, this work presents an efficient\nelectrochemical approach for C4-functionalization of inert pyridines; however, several issues\nrequire clarification before further consideration.\nAuthors’ Response: We sincerely appreciate your positive feedback and constructive suggestions\non our manuscript.\n1. Table 1 shows TEMPO as a reaction component, while Figure 1A-III indicates TEMPO inhibits\nproduct formation. This apparent contradiction requires explanation.\nResponse: Thank you for your question regarding the role of TEMPO in this reaction. To clarify,\nTEMPO acts as a mediator to promote aromatization through oxidation. However, at higher\nconcentrations, it functions as a radical quencher, thereby suppressing the formation of the desired\nproduct. This concentration-dependent dual role of TEMPO has been documented in prior studies\n(Green Chem., 2022, 24, 4928-4934).\n2. In Figure 1A, the authors mentioned that homocoupling occurs when A1 or B1 is reacted\nindividually. However, under standard conditions where both substrates are present simultaneously,\nheterocoupling is exclusively favored with no detectable homocoupling products. This selectivity\npattern requires mechanistic elucidation. The authors are encouraged to perform computational\nstudies to provide a reasonable explanation.\nResponse: Thank you for your suggestions. We would like to clarify that under the standard reaction\nconditions, the system does generate homocoupling products; in fact, a small amount of pinacol\nby-product is also formed during the reaction. Furthermore, when 3 equivalents of 2-phenylpyridine\nand 1 equivalent of 4-methoxyacetophenone were used (Table S1, entry(cid:3031)20), the target product was\nobtained in 57% yield, accompanied by the pyridine homocoupling product in approximately 17%\nyield (yield calculated based on pyridine equivalents). The cross-coupling mechanism in this\nreaction depends mainly on the electronic compatibility between the two distinct radicals, and is\nalso influenced by the rate of radical generation—consistent with our previous reports\n(J.(cid:3031)Am.(cid:3031)Chem.(cid:3031)Soc.(cid:3031)2023,(cid:3031)145,(cid:3031)10967–10973; Org.(cid:3031)Lett.(cid:3031)2025,(cid:3031)27,(cid:3031)4725–4731).\n3. The KIE in Figure 1C is interpreted as evidence for rate-determining C–H cleavage. However,\nfollowing established literature (Angew. Chem. Int. Ed. 2012, 51, 3066), KIE data alone may not\ndefinitively identify the RDS. The text should be revised to acknowledge alternative interpretations.\nResponse: Thank you for your comments. We have revised the expression as follows:“This result\nsuggests that C–H bond cleavage at the pyridyl C4 position is likely involved in the rate-determining\nstep, though the presence of other rate-limiting processes cannot be excluded.” The reference is\ncited as ref 77.\n4. Please clarify: (1) Is \"M06-2x-dd3\" in Figure 2 a typo? (2) Why were different solvation models\n(PCM for optimization, SMD for energies) employed? Reference support for this approach should\nbe provided.\nResponse: (1) Yes. “M06-2x-dd3” in Figure 2 is a typographical error. It has been corrected to M06-\n2X-D3 in the revised figure and associated text.\n(2) We used IEFPCM (PCM) for geometry optimizations and frequency analyses mainly for\nnumerical robustness. Employing SMD during optimization/frequency calculations can\noccasionally make convergence more difficult and may introduce small symmetry-breaking\ndistortions (especially for high-symmetry stationary points), which can lead to spurious\nlow/imaginary frequencies. By contrast, IEFPCM is generally more stable for these tasks, and\nGaussian has specifically improved PCM optimization behavior in recent versions. It is also\ncommonly to use PCM solvent model for optimization and SMD solvent model for high level single\npoint calculation in literatures. (see, Nat. Commun. 16, 10283, (2025); Angew. Chem. Int. Ed. 64,\ne202501924, (2025))\n5. The authors claim “the electron spin predominantly located on the C4 and C6 positions, with little\nspin density at the C2 position.” However, only the spin populations at C2 and C4 are labeled in\nFigure 1. It is suggested to label the spin populations at N and C6 as well.\nResponse: Thank you for the suggestions. We agree that explicitly labeling the spin populations at\nN and C6 would make the statement clearer and improve the readability of figure. In the revised\nmanuscript, we have added the spin population labels at N and C6 and updated the figure\naccordingly.\n)lom/lack(\nygrenE\nevitaleR\nFigure 3\n6. Figure S10D is identical to Figure 1. Please remove this redundant figure.\nResponse: Thank you for your suggestion. The redundant figure has now been removed.\nREVIEWER COMMENTS\nReviewer #1 (Remarks to the Author):\nThe author has addressed all the concerns raised by the reviewers.Therefore, I\nrecommend acceptance of the manuscript in its present form.\nAuthors' Response: Thank you very much for your support in the publication of our\nwork.\nReviewer #2 (Remarks to the Author):\nI appreciate the authors' efforts in addressing the previous concerns. They have added\nnew substrates, provided Faradaic efficiency data, and made adjustments to the title and\nterminology. However, several important issues remain that need clarification before\nthe manuscript meets the standards expected for Nature Communications.\nThe work presents a useful electrochemical method for functionalizing pyridines under\nmild, metal-free conditions. The method works best for 2-aryl substituted pyridines and\ncan be extended to simpler pyridines and other heterocycles, though with lower yields\nand harsher conditions. The mechanistic studies are informative, and the practical\napplications are valuable. My concerns are primarily about presentation and ensuring\nclaims align with the actual data.\nAuthors' Response: We sincerely thank you for your great effort in evaluation of our\nwork and your insightful comments.\nThe following points require attention:\n1. Mismatch Between Title and Narrative\nWhile the title was changed to remove \"C4-\", the introduction and main narrative still\nheavily emphasize C4-functionalization of \"inert pyridines.\" This creates a fundamental\ndisconnect: the title suggests a general method for pyridine hydroxyalkylation, but the\ntext continues to emphasize C4-selectivity that is actually substrate-dependent rather\nthan method-controlled.\nAdditionally, the term \"paired electrolysis\" in the title may overstate the efficiency of\nthe system given the very low Faradaic efficiency (3.86%, see point 4 below), which\nsuggests that paired electrolysis is not the dominant pathway.\nSuggestion: The title and narrative need to be aligned to accurately represent the scope\nand nature of the work. Options include: (a) revise the introduction to match the current\ntitle, clearly stating that the method works best for 2-substituted pyridines where\nsubstrate electronics favor C4-selectivity, and discuss the low FE honestly; OR (b)\nchange the title to more accurately reflect the actual scope and mechanism, such as\n\"Room-Temperature Electrochemical C4-Hydroxyalkylation of 2-Arylpyridines\"\n(removing \"paired electrolysis\" claim and specifying substrate class). The key issue is\nconsistency—the title, narrative, and data should all align to give readers an accurate\nunderstanding of what has been achieved.\nAuthors' Response: Thank you for your constructive suggestions. We have added a\nqualifying statement specifying 2-substituted pyridines when describing the C4-\nregioselectivity of the method to accurately represent the scope and nature of the work,\nand have also included a discussion of Faradic efficiency in the main text. In addition,\nwe have revised \"paired electrolysis\" as “electrochemical method” or “electrochemical\nstrategy” in the manuscript text to align with the reaction outcomes.\n(In addition, we calculated the Faradic efficiency (FE) of this transformation. Under\nstandard reaction, the FE is relatively low (3.86, see Supplementary Table S1). Given\nthat hydrogen evolution is not significant for most substrates, we attribute the low FE\nprimarily to an unproductive cycle, where the radicals generated from the single-\nelectron reduction of substrates are re-oxidized by TEMPO before radical cross-\ncoupling. As product formation necessitates radical cross-coupling following\nsimultaneous single-electron reduction of substrates with high reduction potentials, the\nobserved low FE remains acceptable for reaction conducted in an undivided cell with\nnon-sacrificial anode. Comparable FE values have been reported in similar\nelectrochemical systems involving multiple single-electron transfers of the substrates\nwith high reduction potential.[80])\n2. Scope and \"Inert Pyridines\" Claims\nThe newly added substrates show the limitations more clearly:\n• C46 (pyridine): 40% yield with modified conditions\n• C47 (2-methylpyridine): 24% yield\n• C52' (4-CF₃-pyridine): 29% yield, C2-selective\n• C54' (3-CF₃-pyridine): 31% yield, C2-selective\nThese results indicate the method is optimized for 2-aryl substituted pyridines rather\nthan being generally applicable to \"inert pyridines.\" The DFT calculations support this:\nC4-selectivity arises primarily from substrate electronics (the 2-aryl substituent)\ncombined with steric effects, not from the electrochemical method itself. This is\nsubstrate-controlled selectivity, not method-controlled selectivity.\nSuggestion: Please clarify in the text that the method works best for 2-aryl (or 2-\nheteroaryl) substituted pyridines. The \"inert pyridines\" framing should be softened to\naccurately represent the scope and avoid overstating the generality.\nResponse: Thank you for pointing out this issue. We have softened the wording\nregarding \"inert pyridines\" in the main text and clarify the reaction scope to accurately\nreflect its generality.\n(Although optimal results are obtained with pyridines bearing 2-aryl or 2-electron-with\ndrawing groups, the method is also applicable to 3-substituents, 4-substituents and\npyridine with acceptable yield and regioselectivity.)\n3. Comparative Claims Need Support\nIn the response letter, you state that selectivity \"is in line with, or exceeds\" recent studies\n(citing JACS 2024, 146, 24257 and Nat. Chem. 2021, 13, 1207). However, this\ncomparison does not appear in the manuscript. If your regioselectivity is indeed\nimproved compared to these methods, showing this comparison would strengthen your\nwork. Additionally, explaining why your method achieves better regioselectivity (if\napplicable) would be valuable.\nSuggestion: Either add a brief comparative table or discussion showing how your\nmethod compares to these references, or remove the comparative claim. If you choose\nto include the comparison, please explain the basis for any superiority in regioselectivity.\nResponse: We thank the reviewer for this helpful suggestion to improve the manuscript.\nWe have removed the comparative claim as suggested.\n4. Faradaic Efficiency Discussion Missing\nThe calculated Faradaic efficiencies (3.86% standard, 3.12% scale-up) are quite low,\nindicating that ~96% of electrons are consumed by side reactions rather than product\nformation. This raises significant concerns about the \"high efficiency\" and \"paired\nelectrolysis\" claims. For context, well-functioning paired electrolysis systems typically\nshow FE >30%.\nI understand that low FE can occur in undivided cells with competing H₂/O₂ evolution,\nbut this should be discussed transparently in the main text rather than just reported in\nthe SI. The low FE directly contradicts the \"efficient paired electrolysis\" narrative\npresented throughout the manuscript.\nSuggestion: Add a paragraph discussing the FE values and their implications for the\nefficiency claims. Either explain why low FE is acceptable for this system (e.g.,\noperational simplicity outweighs electron efficiency), or acknowledge it as a limitation\nthat could be improved in future work. Claims about \"high atom economy\" and\n\"efficient paired electrolysis\" should be qualified or removed given the 3.86% FE.\nResponse: We have included a discussion Faradic efficiency in the main text as request,\nand have changed “paired electrolysis” to “electrochemical method” or\n“electrochemical strategy” in the whole manuscript.\n(In addition, we calculated the Faradic efficiency (FE) of this transformation. Under\nstandard reaction, the FE is relatively low (3.86, see Supplementary Table S1). Given\nthat hydrogen evolution is not significant for most substrates, we attribute the low FE\nprimarily to an unproductive cycle, where the radicals generated from the single-\nelectron reduction of substrates are re-oxidized by TEMPO before radical cross-\ncoupling. As product formation necessitates radical cross-coupling following\nsimultaneous single-electron reduction of substrates with high reduction potentials, the\nobserved low FE remains acceptable for reaction conducted in an undivided cell with\nnon-sacrificial anode. Comparable FE values have been reported in similar\nelectrochemical systems involving multiple single-electron transfers of the substrates\nwith high reduction potential.[80])\n5. NMR Spectra Expansion Incomplete\nWhile some NMR spectra now show expanded aromatic regions, many still remain\ncompressed. Please provide systematic expansion of aromatic regions for all products\nto allow verification of structural assignments.\nResponse: Thank you for your suggestion. We have now provided the systematic\nexpansion of aromatic regions for all products.\nOverall Assessment\nThis work demonstrates several valuable features:\n• A metal-free electrochemical approach for pyridine functionalization\n• Room temperature, operationally simple conditions\n• Mechanistic insights through CV, DFT, and control experiments\n• Practical utility for late-stage functionalization of 2-aryl pyridines\n• Air and moisture tolerance\nThese are legitimate contributions. However, the presentation needs substantial\nadjustment to ensure claims accurately reflect the data:\n• Scope: More specific than \"inert pyridines\"—works best for 2-aryl substituted\npyridines, with poorer performance for simpler pyridines\n• Efficiency: Lower than suggested by current claims (3.86% FE needs honest\ndiscussion)\n• Regioselectivity: Substrate-dependent (electronic and steric effects), not method-\ncontrolled\n• Generality: The \"inert pyridine\" framing oversells the actual scope\nNature Communications readers expect rigorous, honest reporting. With appropriate\nrevisions addressing the five points above, this work should be presented as \"an\nelectrochemical method for hydroxyalkylation of 2-aryl substituted pyridines under\nmild, metal-free conditions\" rather than as a general solution for functionalizing \"inert\npyridines\" with \"high efficiency.\"\nIf the authors can address these presentation issues and align their claims with the actual\ndata, the manuscript could make a useful contribution to the electrochemical synthesis\nliterature.\nResponse: Thank you for the above suggestions to improve our manuscript. We have\nrevised the manuscript in accordance with the review's comments to ensure consistency\nbetween the results presented in the main text and the actual work performed.\nReviewer #3 (Remarks to the Author):\nPrevious comments have been properly addressed. This work is recommended for\npublication with Nature Communications.\nResponse: Thank you very much for your support in the publication of our work.\nReviewer #4 (Remarks to the Author):\nThe authors have addressed the questions reasonably well and I recommend the\nmanuscript for publication in Nature Communications.\nResponse: Thank you very much for your support in the publication of our work."}]

Decision letter