In Situ Enzyme-Free Polymerization

• In situ enzyme-free polymerization is a process where polymers form directly at a functional interface without biological catalysts, using reactive substrates and controlled environments.
• This technique is applied across varied systems, including graphene-based conductive films, radical polymerization around nanomaterials, and in situ passivation layers in lithium-ion batteries.
• By achieving precise spatial control, this method enhances device performance and stability while also offering insights into prebiotic and abiotic polymer formation.

In situ enzyme-free polymerization denotes polymer formation directly at the interface, surface, dispersion microenvironment, or geochemical setting where the resulting material is intended to function, without recourse to biological catalysts. Across the literature, the term spans chemically oxidized growth of PEDOT on covalently functionalized graphene, interfacial nylon formation on surfactant-coated graphene, radical polymerization around black phosphorus nanoflakes, HRP-like nanozyme-catalyzed conductive-polymer growth on stretchable electrodes, additive self-polymerization within battery interphases, and even abiotic condensation scenarios invoked for icy moons and prebiotic sequence evolution (Kovaříček et al., 2021, Das et al., 2010, Passaglia et al., 2018, Li et al., 25 Aug 2025, Barua et al., 30 Nov 2025, Kimura et al., 2015, Walker et al., 2012). In this usage, “enzyme-free” excludes biological catalysts but does not imply the absence of oxidants, radical initiators, reactive interfaces, or inorganic/nanozyme catalytic centers.

1. Conceptual scope and defining features

The central feature of in situ enzyme-free polymerization is spatial coincidence between polymer growth and functional deployment. In materials systems, this means that polymerization occurs selectively on grafted graphene, at the surface of dispersed graphene sheets, around exfoliated black phosphorus nanoflakes, on porous Au@Ag nanowire electrodes, or on silicon-anode interphases during idle aging (Kovaříček et al., 2021, Das et al., 2010, Passaglia et al., 2018, Li et al., 25 Aug 2025, Barua et al., 30 Nov 2025). In planetary and prebiotic contexts, “in situ” instead denotes polymerization within the local physicochemical environment itself, such as the shallow icy crust of Europa or periodically hydrated-dehydrated surface lattices in kinetic simulations (Kimura et al., 2015, Walker et al., 2012).

A second defining feature is that localization is achieved by the reaction medium or substrate architecture rather than by enzymes. Covalently tethered phenylsulfonate groups on graphene act as anchor sites and stationary counter-ions for PEDOT growth; surfactant admicelles create a nanometric water-organic interface for nylon formation; bP nanoflakes confine radical propagation and termination; porous Au shells activate $\mathrm{H_2O_2}$ and confine oxidation to conductor surfaces; partially delithiated Si surfaces initiate FEC self-polymerization; and low-temperature thermodynamics or hydration-dehydration cycling define the accessible reaction window in abiotic settings (Kovaříček et al., 2021, Das et al., 2010, Passaglia et al., 2018, Li et al., 25 Aug 2025, Barua et al., 30 Nov 2025, Kimura et al., 2015, Walker et al., 2012).

A concise classification of representative systems is given below.

System	Polymerization drive	In situ locus
EDOT $\rightarrow$ PEDOT on graphene	FeCl$_3$ oxidative chemistry	Covalently functionalized, patterned graphene
Sebacoyl chloride + diamine $\rightarrow$ nylon	Interfacial condensation	Surfactant-coated dispersed graphene
MMA $\rightarrow$ PMMA	AIBN radical polymerization	Around exfoliated bP nanoflakes
ETE-S $\rightarrow$ PETE-S	Au@Ag HRP-like catalysis with $\mathrm{H_2O_2}$	Stretchable Au@Ag/PDMS electrodes
FEC self-polymerization	One-electron reduction at Si surface	Aging silicon-anode SEI
Amino acid / nucleotide condensation	Low-temperature thermodynamics	Shallow ice crust on Europa
Sequence replication model	Dehydration-driven assembly and replication	Hydration-dehydration cycled lattice

This range suggests that the category is mechanistic rather than material-specific. A plausible implication is that the most useful unifying criterion is not polymer class, but reaction localization under non-enzymatic control.

2. Surface-confined polymerization on graphene and related carbon interfaces

A canonical materials example is the in situ oxidative polymerization of 3,4-ethylenedioxythiophene on photolithographically patterned, covalently functionalized graphene. Monolayer graphene is grown by low-pressure CVD on copper and transferred onto $\mathrm{Si/SiO_2}$, after which lithographic patterning defines stripe widths of $3$–$10\,\mu\mathrm{m}$ with gaps as small as $\rightarrow$0. Diazonium chemistry introduces 4-phenylsulfonate groups, with Raman D/G analysis giving a defect density $\rightarrow$1 and $\rightarrow$2. Immersion in EDOT/FeCl$\rightarrow$3 solution then yields polymerization exclusively on the grafted graphene, producing PEDOT:graphene bilayers with AFM thicknesses of $\rightarrow$4–$\rightarrow$5 in acetonitrile and $\rightarrow$6–$\rightarrow$7 in propylene carbonate; Raman mapping shows PEDOT bands at $\rightarrow$8, $\rightarrow$9, $_3$0, and $_3$1 only on functionalized graphene (Kovaříček et al., 2021).

The underlying oxidative sequence is explicitly chemical and enzyme-free. Fe$_3$2 oxidizes EDOT to a radical cation, two $_3$3 units couple at their $_3$4-positions, and chain growth proceeds by repeated oxidation-coupling cycles, while the phenylsulfonate groups tethered to graphene stabilize the positive charge on the PEDOT backbone. The simplified overall reaction is given as

$_3$5

Spectroelectrochemical Raman measurements over $_3$6 to $_3$7 vs. Ag/Ag$_3$8 show graphene G-mode shifts from $_3$9 at $\rightarrow$0 to $\rightarrow$1 at $\rightarrow$2 and $\rightarrow$3 at $\rightarrow$4, with G-mode area increasing $\rightarrow$5 at $\rightarrow$6 and $\rightarrow$7 at $\rightarrow$8. Doping levels of $\rightarrow$9–$\rightarrow$0 were reported, and negative doping is more efficient than in bare graphene (Kovaříček et al., 2021).

Electrical transport in the same system emphasizes that in situ growth can reshape carrier asymmetry. Pristine graphene was p-doped with $\rightarrow$1 and $\rightarrow$2; after diazonium grafting, hole mobility dropped to $\rightarrow$3; after PEDOT growth, highly asymmetric I–V curves gave $\rightarrow$4, $\rightarrow$5, and a Dirac point at $\rightarrow$6 in $\rightarrow$7 at $\rightarrow$8, shifting to $\rightarrow$9 under ambient conditions (Kovaříček et al., 2021). The reported interpretation is that hole transport is strongly enhanced while electron conduction is suppressed by PEDOT’s positive polarity.

A distinct graphene-based route uses interfacial polycondensation rather than oxidation. In dispersed pristine graphene stabilized by $\rightarrow$0 w/v SDBS in water, carbon tetrachloride containing $\rightarrow$1 sebacoyl chloride swells the hydrophobic cores of surfactant admicelles at the graphene surface. Subsequent dropwise addition of hexamethylenediamine drives localized nylon 6,10 formation through the acid-chloride/amine condensation

$\rightarrow$2

AFM shows SDBS/graphene heights of $\rightarrow$3–$\rightarrow$4 and nylon/SDBS/graphene heights of $\rightarrow$5–$\rightarrow$6, consistent with a $\rightarrow$7–$\rightarrow$8 polymer shell. The nylon-coated dispersions remain stable down to pH $\rightarrow$9–$\mathrm{H_2O_2}$0, whereas SDBS/graphene aggregates at pH $\mathrm{H_2O_2}$1, and after freeze-drying the viscosity curve of nylon/SDBS/graphene essentially overlaps the pre-drying trace (Das et al., 2010). This system clarifies that in situ enzyme-free polymerization can be driven by a confined interface even when no explicit catalyst is used.

3. Radical and nanozyme-mediated growth in hybrid nanomaterials and biointerfaces

In situ radical polymerization has also been used as a coupled exfoliation-passivation strategy for black phosphorus. In one formulation, $\mathrm{H_2O_2}$2 of ground bP crystals are sonicated in $\mathrm{H_2O_2}$3–$\mathrm{H_2O_2}$4 MMA within a $\mathrm{H_2O_2}$5 Schlenk tube under $\mathrm{H_2O_2}$6 and ice-bath cooling, using a $\mathrm{H_2O_2}$7 tip at $\mathrm{H_2O_2}$8, $\mathrm{H_2O_2}$9 input, and $\mathrm{Si/SiO_2}$0 amplitude for $\mathrm{Si/SiO_2}$1, giving a hydrodynamic diameter $\mathrm{Si/SiO_2}$2. After a $\mathrm{Si/SiO_2}$3 nitrogen purge, AIBN is added at $\mathrm{Si/SiO_2}$4 relative to monomer, and MMA polymerization proceeds at $\mathrm{Si/SiO_2}$5 for $\mathrm{Si/SiO_2}$6 (Passaglia et al., 2018).

The kinetic description follows standard radical-polymerization notation:

$\mathrm{Si/SiO_2}$7

The reported confinement effect is that growing radicals near bP surfaces have reduced mobility, lowering termination and increasing $\mathrm{Si/SiO_2}$8 and $\mathrm{Si/SiO_2}$9. SEC/GPC data support this: PMMA_bP_C gave $3$0, $3$1, and $3$2, compared with blank PMMA_C values of $3$3, $3$4, and $3$5 (Passaglia et al., 2018). Raman, $3$6 solid-state MAS NMR, and XRD further indicate superior passivation for the in situ route: bP modes remain detectable after $3$7–$3$8 months or $3$9 UV-aging, and PMMA_bP_C retains bP peaks unchanged after $10\,\mu\mathrm{m}$0 months ambient exposure (Passaglia et al., 2018).

In bioelectronics, a different enzyme-free route uses porous Au-coated Ag nanowires with HRP-like catalytic properties. Here, $10\,\mu\mathrm{m}$1 ETE-S in DI water is combined with $10\,\mu\mathrm{m}$2 $10\,\mu\mathrm{m}$3, the pH is adjusted to $10\,\mu\mathrm{m}$4–$10\,\mu\mathrm{m}$5 by HCl addition, and incubation is carried out at $10\,\mu\mathrm{m}$6–$10\,\mu\mathrm{m}$7 for $10\,\mu\mathrm{m}$8, or up to $10\,\mu\mathrm{m}$9 for complete coverage. The mechanistic scheme is written in analogy to HRP’s Fe(III)/Fe(IV) cycle but with Au(0)/Au(I) redox couples: surface-bound oxidizing species generated from $\rightarrow$00 convert ETE-S into $\rightarrow$01, which then couple into oligomers and ultimately a conjugated polymer (Li et al., 25 Aug 2025).

The reported kinetics are pseudo-first-order in $\rightarrow$02 and monomer concentration,

$\rightarrow$03

with film thickness increasing approximately linearly according to

$\rightarrow$04

and surface coverage described by

$\rightarrow$05

SEM and TEM show a conformal shell of PEDOT-like polymer of $\rightarrow$06–$\rightarrow$07 after $\rightarrow$08, with a granular $\rightarrow$09 surface texture atop Au sub-$\rightarrow$10 grains (Li et al., 25 Aug 2025).

The principal bioelectronic outcome is impedance reduction on stretchable neural electrodes. For $\rightarrow$11 microelectrodes, pristine Au@Ag gives $\rightarrow$12, while after PETE-S polymerization $\rightarrow$13, corresponding to an $\rightarrow$14 reduction. A simplified Randles-type circuit,

$\rightarrow$15

was reported with $\rightarrow$16, $\rightarrow$17 reduced by $\rightarrow$18, and $\rightarrow$19 increased by $\rightarrow$20 upon polymerization (Li et al., 25 Aug 2025). The underlying Au@Ag/PDMS conductor maintains $\rightarrow$21 at $\rightarrow$22 strain and remains $\rightarrow$23 up to $\rightarrow$24 uniaxial elongation, with $\rightarrow$25 resistance drift over $\rightarrow$26 cycles at $\rightarrow$27 strain (Li et al., 25 Aug 2025). This indicates that enzyme-free in situ polymerization can be compatible with mechanically compliant neural interfaces.

4. Interphase polymerization in electrochemical energy storage

In lithium-ion systems, in situ enzyme-free polymerization can occur spontaneously during aging rather than during deliberate synthesis. For high-loading Si ($\rightarrow$28) anodes in baseline $\rightarrow$29 LiPF$\rightarrow$30 in EC-EMC electrolyte, the addition of $\rightarrow$31 FEC was reported to induce self-polymerization during idle aging. Over $\rightarrow$32 days of aging, FEC led to a $\rightarrow$33-fold reduction in irreversible capacity loss in Si-LiFePO$\rightarrow$34 full cells, and projected capacity-retention analysis indicated that cells without FEC fall below $\rightarrow$35 of their initial capacity within $\rightarrow$36 days versus $\rightarrow$37 days with FEC (Barua et al., 30 Nov 2025).

The mechanistic sequence begins with one-electron reduction at a partially delithiated Si surface,

$\rightarrow$38

followed by ring opening of the FEC radical anion and propagation in which the alkoxide radical attacks additional FEC molecules. The resulting polymer motif is described as a polycarbonate backbone carrying $\rightarrow$39 groups,

$\rightarrow$40

This chemistry was inferred from post-mortem FTIR and XPS, which showed a unique polycarbonate band at $\rightarrow$41–$\rightarrow$42 after $\rightarrow$43 days OCV aging with $\rightarrow$44 FEC, as well as a C 1s feature at $\rightarrow$45 assigned to C–F polymeric fragments from FEC self-polymerization (Barua et al., 30 Nov 2025).

The electrical manifestation of this polymer-rich SEI was quantified by power-law fits to long-term interphase resistance in symmetric Si$\rightarrow$46Si cells,

$\rightarrow$47

For $\rightarrow$48, the $\rightarrow$49 FEC case followed $\rightarrow$50 with $\rightarrow$51, whereas the $\rightarrow$52 FEC case followed $\rightarrow$53 with $\rightarrow$54. For $\rightarrow$55, the corresponding fits were $\rightarrow$56 and $\rightarrow$57 (Barua et al., 30 Nov 2025). The interpretation reported in the source is that the FEC-free case exhibits mixed transport-reaction growth with significant dissolution, whereas the FEC-containing case shows diffusion-controlled impedance growth suggestive of a robust passivation film.

Electrochemical performance after aging was correspondingly different. Before aging, first-cycle coulombic efficiency over cycles $\rightarrow$58–$\rightarrow$59 at C/10 was $\rightarrow$60 for $\rightarrow$61 FEC and $\rightarrow$62 for $\rightarrow$63 FEC. After $\rightarrow$64 days idle aging, harvested and reassembled full cells showed first-cycle CE of $\rightarrow$65 for $\rightarrow$66 FEC and $\rightarrow$67 for $\rightarrow$68 FEC, with subsequent cycling $\rightarrow$69 in the FEC case (Barua et al., 30 Nov 2025). This suggests that in situ polymerization can function as a passivation-engineering mechanism rather than only as a route to bulk polymer films.

5. Abiotic polymerization in prebiotic and planetary settings

The concept of in situ enzyme-free polymerization is not confined to engineered materials. In thermodynamic analyses of Europa-like icy environments, amino-acid and nucleotide condensation reactions were evaluated using apparent standard molal Gibbs energies propagated with the revised Helgeson–Kirkham–Flowers equations of state. For a general dehydration-condensation reaction,

$\rightarrow$70

with equilibrium constant

$\rightarrow$71

Kimura and Kitadai examined peptide-forming reactions such as $\rightarrow$72 and the nucleoside-forming reaction adenine + ribose $\rightarrow$73 adenosine + $\rightarrow$74 (Kimura et al., 2015).

From the reported $\rightarrow$75 curves, peptide formation becomes exergonic below threshold temperatures of $\rightarrow$76 for glycine dimerization, $\rightarrow$77 for $\rightarrow$78, $\rightarrow$79 for $\rightarrow$80, and $\rightarrow$81 for $\rightarrow$82. For nucleoside formation at $\rightarrow$83, $\rightarrow$84 for $\rightarrow$85. Europa’s observed surface temperatures of $\rightarrow$86 in the equatorial region and $\rightarrow$87 in the polar region, together with steady-state heat-flow models using

$\rightarrow$88

led to the conclusion that only the top few kilometers of the ice crust remain below the threshold for glycine dimerization, while below $\rightarrow$89–$\rightarrow$90 peptide formation becomes endergonic again (Kimura et al., 2015). The explicit limitation is that kinetics were not addressed.

A complementary but kinetic perspective appears in the model of universal sequence replication under hydration-dehydration cycling. There, two monomer types A and B, their precursors pA and pB, and fixed-length polymers of $\rightarrow$91 undergo spontaneous dimer-nucleated assembly, universal template-directed replication, hydrolytic depolymerization, and surface-confined diffusion. The mean-field equations in the hydrated phase include

$\rightarrow$92

with analogous equations for $\rightarrow$93 and each sequence $\rightarrow$94 (Walker et al., 2012). Simulations were performed on a $\rightarrow$95 periodic lattice over typically $\rightarrow$96 cycles, with typical parameters $\rightarrow$97, $\rightarrow$98, $\rightarrow$99, $_3$00 varied $_3$01–$_3$02, and $_3$03 varied $_3$04–$_3$05 (Walker et al., 2012).

Under non-functional conditions with $_3$06, $_3$07, $_3$08, $_3$09, and $_3$10, total polymers were $_3$11, with $_3$12 of monomers sequestered; extant species numbered $_3$13–$_3$14; sequence exploration proceeded at $_3$15–$_3$16 new sequences per cycle; and local diversity $_3$17 was $_3$18–$_3$19 bits/site (Walker et al., 2012). For $_3$20 and $_3$21, stable clusters $_3$22–$_3$23 polymers wide emerged. When one A-zyme was introduced at $_3$24 cycles with $_3$25, its lineage lifetime became $_3$26–$_3$27 longer than that of average nonfunctional species, and the total polymer population rose $_3$28 above the nonfunctional baseline (Walker et al., 2012). Although this work is in silico, it places enzyme-free polymerization within a general framework of reversible assembly, hydrolysis, diffusion, and emergent selection.

6. Cross-cutting mechanisms, analytical signatures, and unresolved issues

Across these systems, localization is achieved by several recurrent mechanisms. One is fixed interfacial functionality: phenylsulfonate groups on graphene both nucleate PEDOT and act as stationary counter-ions (Kovaříček et al., 2021). A second is reaction confinement by mesoscale architecture, as in surfactant-swollen admicelles on graphene or high-surface-area porous Au shells on Ag nanowires (Das et al., 2010, Li et al., 25 Aug 2025). A third is interfacial confinement of radical dynamics, proposed for bP where reduced mobility near nanoflake surfaces lowers termination (Passaglia et al., 2018). A fourth is autonomous surface chemistry, exemplified by FEC self-polymerization at Si during idle aging (Barua et al., 30 Nov 2025). In abiotic models, confinement is effectively environmental: thermodynamic favorability is restricted to the shallow cryosphere on Europa, while kinetic cycling in the prebiotic replication model alternates between polymer-generating and polymer-degrading phases (Kimura et al., 2015, Walker et al., 2012).

Analytical practice is correspondingly multimodal. AFM, SEM, TEM, and Raman establish nanoscale thickness, coverage, and spectral fingerprints for PEDOT/graphene, nylon/graphene, and PETE-S/Au@Ag systems (Kovaříček et al., 2021, Das et al., 2010, Li et al., 25 Aug 2025). XPS and FTIR reveal interphase chemistry in PETE-S-coated electrodes and FEC-derived SEI films, including S 2p peaks at $_3$29 for PEDOT sulfonate doping, a C 1s $_3$30–$_3$31 shake-up at $_3$32 in the nanozyme-grown polymer, and a polycarbonate FTIR band at $_3$33–$_3$34 in aged Si electrodes (Li et al., 25 Aug 2025, Barua et al., 30 Nov 2025). For bP hybrids, Raman, $_3$35 MAS NMR, XRD, DSC, and TGA distinguish passivation efficacy from simple physical mixing: METHOD C retained a bP signal of $_3$36 of total $_3$37 area, while oxidized species were $_3$38, whereas METHODS A and B reached oxidized fractions up to $_3$39 (Passaglia et al., 2018). In the graphene-dispersion work, rheology provided an operational measure of post-lyophilization network recovery (Das et al., 2010).

Several limitations recur. In PEDOT-on-graphene, film thickness remained in the $_3$40–$_3$41 range and finer tuning via reaction time was not observed; the demonstration was limited to small lab-scale wafer chips, residual Fe/Cl contamination was reported at $_3$42, and photolithography limited resolution to a few micrometers (Kovaříček et al., 2021). In the Au@Ag nanozyme system, excess $_3$43 can overoxidize the conductive polymer, the monomer scope was demonstrated only for sulfonated EDOT, and pattern resolution was limited by manual casting (Li et al., 25 Aug 2025). In the bP case, a gradient of flake dispersion remained even though passivation was improved (Passaglia et al., 2018). In the Europa analysis, $_3$44 was explicitly noted to be necessary but not sufficient, since kinetics, catalytic surfaces, and internal heat sources were neglected (Kimura et al., 2015). In the prebiotic replication model, the conclusions depend on an idealized lattice, fixed polymer length, and prescribed rate constants (Walker et al., 2012). These constraints indicate that “enzyme-free” is not a uniform technology but a family of strategies whose performance depends on how localization, reactivity, and stability are jointly engineered.

A common misconception is that enzyme-free polymerization is intrinsically harsh. The literature does not support that generalization. Some implementations indeed rely on strong oxidants such as FeCl$_3$45 (Kovaříček et al., 2021), yet others operate at near-neutral pH and room temperature through nanozyme catalysis (Li et al., 25 Aug 2025), or arise spontaneously during storage without applied bias (Barua et al., 30 Nov 2025). Another misconception is that enzyme-free implies nonselective bulk deposition. The opposite behavior is frequently the design objective: growth can be exclusive to grafted graphene, confined to exposed Au@Ag electrode windows, or localized to surfactant-structured graphene interfaces (Kovaříček et al., 2021, Li et al., 25 Aug 2025, Das et al., 2010). This suggests that the field is defined less by the absence of biology than by the deliberate use of substrate chemistry, interfacial transport, and physicochemical environment to place polymer exactly where function is required.