Marie-Ange Lespinasse

Dr Kaiyao Wei

Dr Justine Perrin

Dr Matthias Winkler

Sieme Hamaidia

Dr Alexis Leroy

Zuzana Macek Jilkova

Christian Philouze

Dr Patrice N Marche

Dr Carlo Petosa

Dr Jérôme Govin

Dr Anouk Emadali

Yung-Sing Wong
email: yung-sing.wong@univ-grenoble-alpes.fr

Y.-S Wong

Dr Z Macek Jilkova

Dr Z Macek Jilkovai

Enantioselective approach for expanding the three-dimensional space of tetrahydroquinoline to develop bet bromodomain inhibitors

Keywords: anti-inflammatory agent, aza-ortho-quinone methide, BET bromodomain inhibitor, organocatalysis, tetrahydroquinoline

ou non, émanant des établissements d'enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

Introduction

The pharmaceutical industry is faced with the challenge of producing large quantities of active pharmaceutical ingredients, often with sophisticated structures. However, chemistry must remain simple and greener to address both production cost and environment impact by focusing on reducing synthetic steps (one-pot, multicomponent reactions), saving atoms, and using catalysis to limit resources and waste. 1 Another challenge for this industry is to selectively produce chirality, which is known to increase clinical success. The presence of chiral centers induces the formation of complex three-dimensional structures that give the drug specificity towards its biological target. This requires the production and purification of a single enantiomer, which can have significant impacts on the cost and environmental burden of their production. 3 Furthermore, in the case of compounds with several asymmetric centers, accessibility to multiple combinations of configurations must remain trivial to simplify the optimization step and, subsequently, the production of the best diastereomer at large scale. In this prospect, we were interested in the synthesis of chiral poly-substituted 1,2,3,4-tetrahydroquinoline (THQ) scaffold which is a widespread pharmacophore group also present in natural products. 4,5 Examples include the natural product Helquinoline, 6 the synthetic products Torcetrapib, 7 MK-8318 8 and compound 1 9 (Figure 1). In all these bioactive products, chirality plays an important role since having the correct enantiomer is essential for bioactivity and selectivity.

Scheme 1. Enantioselective Povarov reactions to give tri-substituted tetrahydroquinoline.

There are many synthetic routes to access chiral THQ, 5 but the simplest and most versatile to implement with a multicomponent approach remains the asymmetric Povarov reaction. 10,11 Synthesis of highly optically pure THQs can be achieved by enantioselective Povarov reactions based on a [4+2] cycloaddition or a Mannich/Friedel-Craft sequence mechanism. 4 The seminal work by Kobayashi and co-workers showed that trisubstituted THQ can be obtained in good enantioselectivity with a cis-2,3 and cis-3,4 configuration (Scheme 1, Eq.1). 12 However, the method was limited to ortho-phenolic substituted imines. More versatile enantioselective approaches use chiral catalysts such as Jacobsen urea (Eq. 2) 13 and phosphoric acid involved in a multicomponent approach described by Zhu, Masson and coworkers (Eq. 3) 14 that yield 2,3,4-substituted THQ with cis-2,3 and cis-2,3 configuration and trans-2,3 and trans-2,3 configuration, respectively. Recently, Chan and co-workers reported the Mannich/Friedel-Craft sequential reaction, catalyzed by the silylated prolinol/POCl3 catalysts, respectively, to give after extraction, the crude transient 4-hydroxyl-THQ 2. The latter must then be oxidized to yield a more stable trans-2,3substituted 2,3-dihydroquinolinone 3 (Eq. 4). 15 The role of the Lewis acid POCl3 was only limited to catalyzing the Friedel-Craft reaction. However, an enantioselective reaction capable of accessing the cis-2,3 and trans-3,4 configuration of 2,3,4substituted THQ has not yet been reported and will expand the three-dimensional space of THQ. Given the potential of THQ to easily generate chemical libraries, 16,17 this would be of great interest to access this rare configuration and open the door to the exploration of new chemical spaces. Scheme 2. One-pot multicomponent reaction including the generation of an aza-ortho-quinone methide species to form cis-2,3 trans-3,4 trisubstituted THQ.

To achieve this goal and keep the process simple, the development of a one-pot multicomponent reaction based on the Mannich/Friedel-Craft reaction sequence was undertaken to incorporate the additional generation of an aza-ortho-quinone methide species 18 simultaneously (Scheme 2). Indeed, the first cis-2,3 configuration can be controlled by the proline catalyzed Mannich reaction. 19 We identified mild conditions to achieve the Friedel-Craft reaction and the subsequent formation of the highly reactive aza-ortho-quinone methide intermediate. The addition of various nucleophiles is tolerated and selectively gives the trans-3,4 configuration. The use of proline also has the advantage of being the simplest and most environmentally begnin organocatalyst. 20 This new configuration has proven to be essential in our program to develop new inhibitors acting on epigenetic mechanisms.

Results and Discussion

The first enantioselective Mannich reaction step used the proline-catalyzed List-Barbas reaction, starting with imine 4 and aldehydes to give the Mannich product 6 with syn selectivity (Scheme 3). 19 At this stage, product 6 does not spontaneously cyclize to form 4-hydroxy THQ 7 and the assistance of a Lewis acid like POCl 3 15 is thus required. Interestingly, o-aminobenzyl alcohols are known to be a precursor able to generate aza-ortho-quinone methide in situ by means of Lewis acid catalysis. 18 Indeed, Lau and coworkers described the use of BF3.OEt2 to catalyze this conversion. 21 In our approach, we wondered whether BF3.OEt2 can both catalyze the Friedel-Craft reaction leading to 7, as well as inducing subsequently the generation of aza-ortho-quinone methide species 8 in the same vessel.

(R)-(-)-1,1'- binaphthyl-2,2'-diyl hydrogenphosphate 50 2 0 -
To test this hypothesis, our initial trial used the same conditions described by Barbas and co-workers, i.e. stirring imine 4 with heptanal 5 in the presence of L-proline (10 mol %) and in dioxane for 24 h at room temperature, to first produce the Mannich product 6 with high control of enantioand diastereoselectivities. 19 We subsequently added to this vessel TMSN 3 (1.3 equivalent) as nucleophile followed by BF3.OEt2 (50 mol%). As expected, the desired THQ 9 was formed in 46% yield and in 89 % ee (Table 1, entry 1).

During the Mannich reaction, we noticed traces of imine 4

remaining after 24 h. When adding BF3.OEt2, this residual material can react with the aldehyde 5 and TMSN 3 to directly generate racemic Povarov product 9. By stirring the Mannich reaction over 48 h to ensure the consumption of all the imine 4, we were able to improve both the yield and the enantiomeric excess (entry 2). Finally, the best condition was to use 15 mol% of proline for 48 h, followed by the addition of 50 mol% of BF3.OEt2 in the presence of the nucleophile (entry 3). Adding more BF3.OEt2 did not improve the yield whereas using a lower amount resulted in incomplete conversion to form the THQ. BF3.OEt2 was shown to be the best catalyst. In this one-pot process, POCl3 used by Chan and co-workers 15 could not yield the desired compound 9 suggesting its inability to generate the aza-ortho-quinone methide species 8 (entries 5 and 6). The use of other Lewis acid catalysts resulted in a slow reaction with low yields (entries 7-9). Finally, we tested whether phosphoric acid as Brønsted acid can trigger the Friedel-Craft/aza-ortho-quinone methide sequence by replacing BF3.OEt2 with (R)-(-)-1,1'-binaphthyl-2,2'-diyl hydrogenphosphate (entry 11). Even in the presence of 50 mol%, the Mannich product 6 remained unchanged. Having the optimized condition in hand, we explored the use of different nucleophiles, aldehydes and imines (Scheme 4). The reaction exhibited good tolerance with various nucleophiles.

Phenyl-heteroatom compounds like phenylthiol, phenylselenium, and aniline were nucleophilic enough to yield the corresponding THQs. In the case of aniline, a major compound 12a was obtained along with a small amount of another minor diastereomer 12b. The alkyl thiols and alkyl alcohols were also successfully grafted on the THQ scaffold (13-16, 22, 23). In contrast, alkyl amines did not yield any desired THQ.

Scheme 4. Enantioselective one-pot synthesis of various tri-substituted THQ; a water was added as nucleophile.

Use of other aldehydes and imines resulted in the corresponding THQ (19-23). Compound 21 was crystallin and its structure was solved by X-ray analysis confirming the cis-2,3 and trans-3,4 configuration (Fig. S1). The hydroxyl THQ 17 was obtained, albeit in low yield, by adding water to the media.

We chose to apply this methodology to the development of inhibitors selective for a specific subset of bromodomains (BDs). 22 The BET (Bromodomain and Extra Terminal) protein family consists of four members (BRD2, BRD3, BRD4 and BRDT) and plays a key role in the regulation of gene transcription. Each member has two bromodomains, BD1 and BD2, which bind to the acetylated lysines of histones and recruit the molecular machinery required to activate transcription. Previous studies have identified numerous BET bromodomain inhibitors, starting with JQ1 and I-BET. 23 Recently, the identification of molecules that selectively inhibit BET BD2, but not BD1, has elicited great interest as they impair the activation of pro-inflammatory genes without affecting the expression of housekeeping and maintenance genes, thereby limiting their toxicity and improving the drug safety profile. 24 Selective BD2 inhibitors have been shown to present a more selective phenotypic fingerprint through the inhibition of key pro-inflammatory mediators. 24 We chose to apply our approach to obtain two diastereomeric THQ structures inspired by compound 1 (Scheme 1) and investigated whether the new accessible configuration would play an important role. Self-Mannich product can be obtained by directly reacting an aniline with two alkyl aldehydes to give the corresponding Mannich products in an enantioselective manner either with a cis-2,3 or a trans-2,3 configuration through proline 19 or silylated prolinol 15 catalysis, respectively. D-proline was used as the catalyst to set up the appropriate bioactive (S)-configuration at position 2 of the THQ. 25 The Mannich reaction was optimized to accommodate the weak nucleophilic aniline 24 (Scheme 5). Indeed, the reaction with aldehyde 25 proved to be slow and cooling to low temperature was essential to maintain good enantioselectivity. Co-formation of the unwanted aldol product also complicated the progress of the reaction as it tends to neutralize the proline catalyst. 26 Finally, performing the reaction at -22°C for 3 days with 30 mol% of D-proline under solvent-free conditions 26 accelerated the reaction while minimizing aldol product formation. Subsequent additions of solvent (EtOAc), followed by the addition of TMSN3 (1.5 equivalent) and BF3.OEt2 (70 mol%) at 0°C afforded THQ 26 in 68% yield and in 91.8 % ee. The reaction was run at larger scale (with 3 g of 24). At this scale and to accelerate the Friedel-Craft reaction and the formation of aza-ortho-quinone methide, more BF3.OEt2 was added (1.36 equivalent). The desired product 26 was isolated in 77.5% yield and in 86.1% ee.

After recrystallization, THQ 26 was obtained in enantiopure form (>99% ee) in 40% yield from aniline 24. THQ 26 was next acetylated to obtain compound 27 on which a simple triazole-phenyl group was installed by Huisgen copper-catalyzed click reaction to lead to the new analogue 28. To make the corresponding diastereomer with the trans-2,3 and trans-3,4 configuration, aniline 24 and propionaldehyde 25 were reacted with silylated prolinol 29 (20 mol%) in MTBE for 2 days at -10°C. 15 Subsequently addition of TMSN3 (1.5 equivalent) and BF3.OEt2 (2.1 equiv.) in the same vessel gave the product 30 with the desired configuration in 59% yield and with 96% ee. Compound 30 was acetylated to give 31. Likely due to steric hindrance around the azido group, the conventional conditions for the copper-catalyzed Huisgen reaction gave less than 40% yield for product 32. By performing the reaction under thermal conditions and in the presence of Cu(CH3CN)4.BF4 catalyst, 27 compound 32 was finally obtained in good yield (92%).

Scheme 5. Diastereodivergent synthesis of bromodomain inhibitors in three steps

The bioactivities of these compounds were evaluated in vitro on recombinant BET Brd4 BD1 and BD2 using a homogeneous time resolved fluorescence (HTRF) assay 28 (Table 2 and see Fig. S2 for IC50 curves). In short, this assay evaluates the ability of compounds to inhibit the interaction of a BD with its natural cognate ligand, here a peptide mimicking the N-terminal tail of histone H4, acetylated on lysine residues 5, 8, 12 and 16. JQ1, a nonselective BD1/BD2 inhibitor, and RVX-208, 29 a selective inhibitor for BD2 versus BD1, were used as controls. The simplest compound 27 showed a significant BET inhibition, with high selectivity (>60-fold) for BD2 versus BD1 (entry 3). The corresponding enantiomer (ent)-27 showed no activity (entry 4) which is consistent with the importance of having an (S)-configuration at position 2 of the THQ. 25 The importance of having the right configuration is highlighted by the lack of or very low inhibitory activity of the compound 31 on BD1 and BD2, demonstrating that the cis-2,3 and trans-3,4 THQ configuration is critical for efficient BET bromodomain inhibition with this scaffold (entry 5). This was also evident for the phenyl substituted analogue 28 (entry 6) compared to its nearly inactive diastereomer 32 (entry 7). Compared to product 27, the presence of the phenyltriazole moiety at position 4 in the compound 28 improved activity approximately 3-fold without significantly altering selectivity for BD2 versus BD1 (entry 6).

Compounds 27, 28, 31, 32 and JQ1 and RVX-208 were next evaluated for their potency as anti-inflammatory agents and for their cytotoxicity on macrophage cells (Figure 2). These compounds led to a dose-dependent decrease of proinflammatory molecules such as IL-6 (Figure 2A) and TNFα (Figure 2B) upon an LPS-mediated induction of inflammation.

Compounds 27 and especially 28 efficiently inhibited the secretion of both molecules, whereas their respective trans-2,3 and trans-3,4 isomers 31 and 32 had no or very weak effects. As previously reported, JQ1 showed cytotoxicity. [START_REF] Mertz | Proc. Natl. Acad. Sci[END_REF] In contrast, all BD2-selective compounds displayed much lower cytotoxicity (Figure 2C). Strikingly, our compounds were significantly less cytotoxic compared to the BD2-selective reference molecule RVX-208. Taken together, compound 28 combines the best efficacy and cytotoxicity profile of all products tested (Figure 2D).

Conclusion

The development of enantiopure bioactive compounds is a challenge for the pharmaceutical industry considering its cost and the subsequent large-scale production. In addition, access to new homochiral configurations opens the door to the exploration of new interaction properties with biological targets. Here we report a one-pot multicomponent reaction that produces THQ via the mild formation of aza-orthoquinone methide species. Coupled with a previous proline catalysis, this protocol provided access to a new configuration allowing for intensification of the structural complexity and diversification from very simple starting materials and catalysts. In our research program on epigenetic inhibitors, the optically pure THQ 26 was easily obtained in large quantities as a building block for the development of BET bromodomain inhibitor. In only two synthetic steps from 26, we were able to identify compound 28 as a selective inhibitor for Brd4 BD2 versus BD1, and as an anti-inflammatory agent with a favourable efficacy/toxicity profile. In this context, the new cis-2,3 and trans-3,4 THQ configuration was critically important for these properties compared to the trans-2,3 and trans-3,4 configuration obtained from previous synthetic methods.

Modulations are underway to improve the pharmacological properties of this lead compound. Given the above results, this simple and cost-effective multicomponent synthesis should open new perspectives for the use of enantiopure THQ as a versatile synthon and as an advanced active pharmaceutical ingredient to enable the production of future drugs.

Experimental Section

Synthesis of (2S,3S,4R)-Ethyl-4-azido-6-methoxy-3pentyl-1,2,3,4-tetrahydroquinoline-2-carboxylate (9) To a solution of ethyl (E)-2-((4methoxyphenyl)imino)acetate 4 (300.0 mg, 1.45 mmol) and freshly distilled heptanal (248.0 mg, 2.17 mmol) in anhydrous dioxane (10 mL) was added L-proline (26.4 mg, 0.21 mmol) at rt. The reaction mixture was stirred for 48 h at the same temperature. After the reaction was complete, TMSN3 (216.0 mg, 1.88 mmol) and BF3.Et2O (90 µL, 0.73 mmol) were added and the mixture was stirred at rt for 1 h. The reaction was quenched with NaOH (2 N) until neutralization, extracted with EtOAc, dried by MgSO4, filtered off and concentrated in vacuo. After flash chromatography (silica gel, 5% EtOAc/cyclohexane), the product 9 was isolated as an oil (333.0 mg, 66.4%); Rf = 0.27 (10% EtOAc/cyclohexane); [α]D 20 +126.

2

 2

Figure 1 .

 1 Figure 1. Various bioactive natural and synthetic 1,2,3,4-tetrahydroquinolines.

Figure 2 .

 2 Figure 2. Evaluation of anti-inflammatory activity and cellular toxicity in J774 macrophage cells. A-B. Level of secreted IL-6 (A) and TNF-α (B) measured after treatment of J774 macrophages with LPS (inflammation inducer) and increasing doses (1-10-100 µM) of BET inhibitor (represented by colour gradient). Data are normalised to LPS-treated conditions. C. Dose-response curves showing impact of BET inhibitor treatment on cellular toxicity evaluated through MTT assay. D. Bubble plot representing anti-inflammatory activity (AI score) versus toxicity (MTT log IC50).

  6 (c 1.03 in CHCl3); IR νmax (thin film, CH2Cl2) 3393, 2954, 2927, 2091, 1734, 1622, 1501, 1463, 1224 cm -1 ; UV : 208, 245, 325; 1 H NMR (400 MHz, CDCl3) δ ppm 0.84 (t, J = 6.9 Hz, 3H), 0.94-1.29 (m, 8H), 1.32 (t, J = 7.1 Hz, 3H), 2.24 (m, 1H), 3.76 (s, 3H), 4.19 (m, 1H), 4.26 (m, 1H), 4.29 (m, 1H), 4.40 (d, J = 2.4 Hz, 1H), 6.64 (d, J = 8.8 Hz, 1H), 6.68 (d, J = 2.8 Hz, 1H), 6.81 (dd, J = 8.8, 2.8 Hz, 1H); 13 C NMR (100 MHz, CDCl3) δ ppm 14.1 (CH3), 14.4 (CH3), 22.6 (CH2), 26.4 (CH2), 27.0 (CH2), 31.8 (CH2), 38.6 (CH), 53.5 (CH), 56.0 (CH3), 61.4 (CH), 61.6 (CH2), 115.6 (CH), 115.8 (C), 116.3 (CH), 117.1 (CH), 136.8 (C), 151.9 (C), 172.5 (C); LRMS (ESI+) m/z (%) 347 (20) [M+H] + , 304 (100); HRMS (ESI+) m/z calc. for C18H27N4O3 347.2078, found 347.2077. Deposition Number (https://www.ccdc.cam.ac.uk/services/structures?id=doi:10. 1002/chem.202202293) 2143668 (for 21) contains the supplementary crystallographic data for this paper. These data are provided free of charge by the joint Cambridge Crystallographic Data Centre and Fachinformationszentrum Karlsruhe http://www.ccdc.cam.ac.uk/structures, Access Structures service. For other experimental Procedures/Data, see Supporting Information

Scheme 3 .

 3 One-pot multicomponent synthesis of enantio-enriched THQ 9 with cis-2,3 and trans-4,4 configuration

Table 1 .

 1 conditions for the THQ 9 synthesis (in dioxane)

	Entry	L-Pro	t1	Cat. 2	Quantity	t2	Yield	ee
		(mol%)	(h)		Cat.2	(h)	of 9
					(mol%)		(%)
	1	10	24	BF3.OEt2	50	1	46	88.9
	2	10	48	BF3.OEt2	50	1	65	92.4
	3	15	48	BF3.OEt2	50	1	66	96.4
	4	20	48	BF3.OEt2	50	1	66	94.4
	5	15	48	POCl3	50	1	0	-
	6	15	48	POCl3	50	24	0	-
	7	15	48	Yb(OTf)3	10	24	33	95.4
	8	15	48	Y(OTf)3	10	24	12	95.4
	9	15	48	Zn(OTf)3	10	24	24	95.3
	10	15	48	InCl3	10	24	24	-
	11	15	48

Table 2 .

 2 Evaluation of bromodomain inhibitors using the HTRF assay for efficacy and BD2/BD1 selectivity.

	Entry	Compound	BD1	BD2	Selectivity
			IC50 (nM)	IC50 (nM)	BD2/BD1
	1	JQ1	67.5	33.6	2
	2	RVX208	30 600	254	150
	3	27	95 300	1510	63
	4	(ent)-27	> 500 000	>50 000	-
	5	31	>400 000	>100 000	-
	6	28	26 100	481	54
	7	32	>100 000	29 900	-

Acknowledgements

This work was supported by the «Agence Nationale de la Recherche» (ANR) under the program Labex Arcane and CBH-EUR-GS, grant number ANR-17-EURE-003 and the ANR, grant number ANR-18-CE18-007. We also acknowledge support from « La Ligue Contre le Cancer-Comité regional de l'Isère », grant number LNCC-R20015CC and « Fond AGIR pour les Maladies Chroniques ». Technical support from the NanoBio Institut de Chimie Moléculaire de Grenoble (NanoBio ICMG, UAR 2607) and the staffs (Marie-Carmen Molina, Isabelle Jeacomine, Nicolas Altounian, Romain Sanahuges, Laure Fort, Amélie Durand and Rodolphe Gueret) are acknowledged for analysis and characterization facilities. We are grateful to the French Alternative Energies and Atomic Energy Commission (CEA) for a PhD studentship (KW).

Entry for the Table of Contents

Simple and selective. Assembling simple functional groups by proline organocatalysis coupled with the addition of BF3.OEt2 in the same flask allows for the mild formation of aza-ortho-quinone methide species that undergo nucleophilic addition to give access to enantio-enriched tri-substituted tetrahydroquinolines with a novel configuration. This advanced three-dimensional structure, after only two additional steps, gave rise to a valuable enantiopure and selective anti-inflammatory compound.

Institute and/or researcher Twitter usernames: dpm_lab


Jian Lv

Lu Yin

Xiaohong Chen

Itthipon Jeerapan

Cristian A Silva

Yang Li

Minh Le

Zhihua Lin

Luwen Wang

Alexander Trifonov

Sheng Xu

Serge Cosnier
email: serge.cosnier@univ-grenoble-alpes.fr

Joseph Wang
email: josephwang@ucsd.edu

Wearable Biosupercapacitor: Harvesting and Storing Energy from Sweat

Keywords: biosupercapacitor, wearable electronics, energy harvesting, stretchable electronics, self-charging

This work demonstrates the first example of sweat-based wearable and stretchable biosupercapacitors, capable of generating high-power pulses from human activity. The allprinted, dual-functional, conformal biosupercapacitor platform can harvest and store energy from sweat lactate. By integrating energy harvesting and storage functionalities on the same footprint of a single epidermal device, the new wearable energy system can deliver high-power pulses and be rapidly self-charged by bioenergy conversion of sweat lactate generated from human activity while simplifying the design and fabrication. The mechanical robustness and conformability of the device are realized through island-bridge patterns and strain-enduring inks. The enhanced capacitance of the biosupercapacitor is realized by the synergistic effect of carbon nanotube ink with electrodeposited polypyrrole on the anode and of porous cauliflower-like platinum on the cathode. In the presence of lactate, the biosupercapacitor shows high power in pulsed output and stable cycling performance. Furthermore, the wearable device can store energy and deliver highpower pulses long after the perspiration stopped. The self-charging hybrid wearable device obtained high power of 1.7 mW cm -2 in vitro, and 343 µW cm -2 on the body during exercise, suggesting considerable potential as a power source for the next generation of wearable electronics.

Introduction

In recent years, the field of wearable electronics has been experiencing a transition from rigid, bulky, and non-conformal devices to miniaturized, flexible, and stretchable devices that conform and adapt to the human body with high intimacy. [1,2] Built upon the work of numerous researchers, the state-of-the-art flexible and stretchable electronics, ranging from mechanical and electrochemical sensors [3][4][5] to complex systems such as multi-sensor arrays, transistor arrays, or electronic displays, [6][7][8][9] have been assembled into a comprehensive ecosystem of various class of devices with distinct functionalities. However, one of the main challenges that impede such wide adoption of flexible and stretchable wearable electronics is the lack of an equally flexible, stretchable, and miniaturized energy source that can supply sufficient power over extended periods. Depending on the functionalities of the device, [10] most wearable electronics consume power ranging from as low as nano-watts to more common milli-watts, with expected usage ranging from a few hours to multiple days. Tremendous efforts have been dedicated to the fabrication of wearable systems to harvest and store energy from various sources, such as human movement, perspiration, sunlight, or the air. [11][12][13][14][15] Among these, wearable biofuel cells (BFCs) that convert chemical energy from human perspiration to electrical energy via enzymatic electrochemical reactions are considered a promising potential solution. [16][17][18][19][20] Nevertheless, one of the key shortcomings of such sweat-based energy harvesters is the need for the constant supply of fuel from human perspiration and the presence of the oxidizer. To address this issue, hybrid devices that combine energy harvesting and storage modules have been reported. [21,22] For example, we demonstrated previously a stretchable textile-based wearable hybrid BFCsupercapacitor (SC) device, where energy is generated from human sweat by the BFC and stored in the SC for later use. [21] However, these integrated hybrid devices still rely on the external connection of the two separate harvesting and storage modules. Some recent efforts demonstrated the possibility of endowing multiple functionalities onto one single device via material innovations, storing energy in multiple modes, or simultaneously harvesting and storing mechanical and chemical energy; however, such multi-functional materials and designs have not been demonstrated for wearable energy systems. [23][24][25] We envision the integration of harvesting and energy storage functionalities at the material level would be more beneficial for the miniaturization of a sustainable high-power energy supply unit for wearable electronics.

Self-charging biosupercapacitors (BSCs) that can store energy and be self-charged via chemical or solar energy conversion have recently attracted considerable attention. [26][27][28] BSCs are biomaterial-based two-electrode systems that combine both BFC and SC functionalities in a single device. [29] Compared with traditional BFCs, the BSCs can generate higher power pulses due to their high-capacitance electrodes that allow rapid discharges. Supercapacitance can be realized by using nanostructured electrode materials to increase electrochemical double-layer capacitance [25] or by the addition of redox molecules [30] and conductive polymers [31] to endow pseudocapacitive behavior. Compared with traditional SCs, which require external power input, self-charging BSCs rely on biocatalysts, such as microbes, organelles, or enzymes, immobilized onto highly capacitive material that allows energy conversion from body fluids or sunlight and its subsequent storage. [29] The dual-functionality feature of BSCs can greatly simplify the design of wearable systems as no external connection is needed, hence improving the energy efficiency and facilitating the miniaturization of the electronic device. For chemical-energy-based BSCs, glucose-and other fuel-based BSCs have been studied. [27] However, to the best of our knowledge, there are no reports on sweat-operated BSCs for on-body wearable applications. For a wearable BSC, the flexibility, stretchability, and skin conformity of the device are of considerable importance. Different materials have been used previously to fabricate electrodes for BSCs, including carbon paper, carbon nanotube (CNT) pellets, or graphite foil. [17,25,30,31] However, these materials are either bulky, rigid, or fragile, and therefore cannot serve as suitable candidates for stretchable conformal wearable electronic devices.

In this work, we demonstrate the first example of an all-printed dual-functional stretchable and wearable BSC, fabricated on top of low-elastic modulus and adhesive elastic films, to harvest and store energy from sweat while maintaining intimate contact with the human skin. This wearable hybrid device, functioning as both a BFC and an SC, is demonstrated to deliver high-power pulses and be rapidly self-charged using enzymatic oxidation of lactate biofuel from human perspiration. Compared to an early multi-module BFC and SC system, [21] this work enabled material-level integration of both functionalities on the same set of electrodes, thus reducing the system complexity and minimizing the device footprint. A scalable and lowcost screen-printing process, using stress-enduring inks, is used to fabricate the BSC (combining the BFC and SC functions on the same printed anode and cathode) and meeting the softness, stretchability, and scalability requirements for this skin-worn wearable device. The stable, conformal, and stretchable energy harvesting-storage dual-function device has been realized through the highly efficient planar electrode design with an island-bridge structure and the use of such strain-enduring inks. [2,32] The island-bridge structure is fabricated on a soft silicone rubber substrate; a non-elastic backbone layer is printed at the bottom of the active electrode "islands" for mechanical support, while silver interconnections are printed in serpentine patterns as "bridges". [33] The stress in the electrode area is thereby distributed to the surrounding flexible region when the device is under deformation, hence ensuring attractive mechanical resiliency.

Compared to the common lactate/O 2 BFC, the greatly enhanced capacitance of the BSC is achieved by the synergistic effect of the CNT-based ink and the electrodeposited polypyrrole conductive polymer on the anode and of the high-surface-area cauliflower-like structured porous Pt on the cathode. The resulting wearable dual-functional BSC can support high-power pulsed output while maintaining stable charge-discharge performance in human sweat-level lactate. The self-rechargeable hybrid device has been mounted on the user's arm and demonstrated harvesting high power up to 1.7 mW cm -2 , suggesting its considerable potential as a power source for the next generation of wearable devices.

Results and discussion

The new soft, dual functional, and skin-worn BSC, integrated on a conformal elastomeric substrate, was designed in a planar structure to harvest and store energy from human sweat (Figure 1A and1B). The surface and structure of the device have been tailored to merge the BFC and SC functions into one single wearable device (Figure 1C). The surface of both electrodes was thus modified to combine the energy harvesting capability with the energy storage functionality. Polypyrrole (PPy) is an attractive electrode material for SC operation owing to its favorable conductivity and pseudocapacitance. [34,35] Therefore, the anode was assembled based on the PPy-electrodeposited multiwalled carbon nanotubes (CNTs) and decorated by lactate oxidase (LOx) enzyme, mediated by 1,4-naphthoquinone (NQ) relay units.

In the anode, the LOx catalyzed lactate oxidation is facilitated by the NQ that regenerates the flavine mononucleotide (FMN) cofactor, allowing continuous fuel oxidation. The cathode is composed of the same CNTs-based electrode, functionalized by an electrodeposited highsurface-area porous Pt catalytic layer, offering high electrochemical double-layer capacitance.

The Pt-catalyzed oxygen reduction reaction provides a driving force for the continuous electron flow from the anode to the cathode, producing and storing electric energy on the same device.

A phosphate buffer solution-polyvinyl alcohol (PBS-PVA) hydrogel was implemented as the electrolyte that allowed free diffusion of sweat to the electrode surface. [22] By such design, the wearable BSC can be continuously charged in open circuit mode, then generating stable high-power pulses on the order of seconds when discharged at a high current (Figure 1D-F).

Screen printing, a low-cost and large-scale fabrication method, was used to manufacture the stretchable and conformal BSC device (Figure S1). The design of the device included an "island-bridge" configuration and was fabricated with customized strain-enduring inks. The "island-bridge" structure is designed to maximize the mechanical robustness of the device, where the functional electrodes are separated into several individual compartments as "islands" and are interconnected by highly stretchable conductive silver interconnections which act as the "bridges". Besides, a flexible, non-stretchable "skeleton" layer with high elastic modulus and mechanical strength is added below the "islands" as reinforcement to further minimize any strain applied to the electrodes. [33] As a result, the strain induced by external deformation can be effectively dispersed around the electrode "islands", and any adverse effects yielded from the deformation can be minimized. Adopted from our previous work, the CNTs and silver inks used in the printing of the stretchable electrodes were customized to obtain outstanding flexibility and stretchability. [36] The stretchability of the CNT ink is inherited from its elastic polyurethane (PU) binder, while the highly conductive and flexible silver ink is made of silver flakes and an elastomeric copolymer binder (polystyrene-block-polyisoprene-block-polystyrene; SIS) [21,36] . Working in conjunction, the elaborate "island-bridge" structure design and ink formulation endow the printed electrodes with electrical conductivity and stability during mechanical deformation. Pt and LOx as catalysts for oxygen reduction and lactate oxidation, with a catalytic onset potential of E ≈ 0.33 V and -0.15 V vs. Ag/AgCl and improved the electrode's capacitance, respectively. The choice of the PPy, as a capacitive polymer, imparts a porous electrode morphology with increased surface area and pseudocapacitance along with an overall stable positive charge in the range of 4-8 pH, which is within the range of the pH of human perspiration. [37] The protonation/deprotonation of nitrogen atoms of the polymer is a primary reason for pseudocapacitive behavior and the ability to store the charge. Evidently, on the bioanode, a deposited porous PPy film has allowed stable LOx immobilization on the carbonaceous electrode matrix and significantly improved the electrode's capacitance (by ~2 folds) (Figure S5).

The energy harvesting performance of the BFC, consisting of the LOx/NQ/PPy-based bioanode and the porous Pt-based cathode, was characterized. Figure 2D shows the power density of BFC when discharged against variable resistances in the presence of different lactate levels. The data plots indicate a typical power curve behavior, showing an increase in opencircuit voltage (OCV) from E = 0.28 V to E = 0.55 V upon elevating the lactate concentration from 0 to 15 mM. The power values were also enhanced upon increasing the lactate level until reaching saturation at 10 mM lactate, with a maximal power density of 80 μW cm -2 at E = 0.38 V. Over the range of 0-5 mM lactate, the power increases linearly with the fuel concentration, demonstrating an increase of ~ 9 μW cm -2 mM -1 . A maximum OCV of E = 0.55 V and the closed-circuit current density of 460 μA cm-2 were reached at 10 mM lactate (Figure 2E). Furthermore, a long-term stability test of the BFC, shown in Figure 2F, demonstrates a stable output with a small standard deviation of 6.4% during the 8 h study period, and effective output of around 100 μW cm -2 at 0.38 V. It should be noted that the OCV established by the cell exceeds the theoretical value of 0.48 V calculated based on electrochemical characterization of the LOx anode and Pt cathode (Figure S3-S4). This effect is attributed to the presence of the redox-active PPy with a charge-storing pseudocapacitance. [38] As the charging of the BSC device increases significantly at higher voltages, the OCV of 0.55 V, established within 2 h, was considered as a practical OCV value.

The capacitance of the BSC can be maximized through several strategies. A CNT-based composite ink was chosen for its attractive active surface area, which greatly enhances the double-layer capacitance of the electrodes, based on our early studies. [21,39] PPy is electropolymerized onto the anode from a pyrrole-containing KClO 4 solution to endow higher capacitance to the bioanode. Pt is electroplated onto the cathode at E = -0.95 V vs. Ag/AgCl to facilitate oxygen-reduction reaction (ORR). Such deposition results in a cauliflower-like structured, high-surface-area Pt, which further enhances the double-layer capacitance of the BSC.

Cyclic voltammetry (CV), used for characterization, demonstrates the significant improvement in BSC capacitance after the PPy and Pt functionalization of the anodes and cathodes, respectively (Figure 2G). LOx is thereafter drop-cast onto the anode and immobilized with chitosan. The effect of the capacitive behavior of the BSC was further validated via the galvanostatic chargedischarge (GCD) technique with different current densities. In Figure 2H, GCD was conducted between 0 to 0.6 V, and the samples with and without PPy and Pt functionalization were tested at a discharge rate of 0.1 mA cm -2 to confirm the increase in capacitance of such treatments. The obtained highest areal capacitance and areal power density of the BSC were calculated to be 27.2 mF cm -2 and 0.22 mW cm -2 , respectively (~2.4 F g -1 and ~20 mW g -1 for the sum of active electrode material weighed ~11 mg cm -2 ), which are considered very advantageous compared to reported BSC literature as shown in Table S1. To determine the self-discharge rate of the BSC, its electrical potential was continuously monitored in the absence of lactate, demonstrating the ability to maintain a voltage above 0.4 V for over 100 min after charging to 0.55 V (Figure S8). The ability of the dual-functional BSC to deliver high-power pulses was evaluated using current-pulse chronopotentiometry (CP). The BSC was discharged at 1 mA cm -2 for 10 ms and left to recharge for 1 min in the absence and presence of 10 mM lactate. The results shown in Figure 3A indicate that in the absence of fuel, the OCV of the BSC is only 0.16 V, and a small power density of 0.13 mW cm -2 is delivered. In contrast, in the presence of 10 mM lactate, the OCV and pulse power increase to 0.55 V and 0.5 mW cm -2 , respectively, indicating the indispensable role of lactate for self-charging BSC.

The performance of the hybrid device was evaluated by applying different current density 10 ms-pulses, ranging from 1 to 10 mA cm -2 . As shown in Figure 3B, after applying the short current pulses, the BSC can deliver power using the charges stored in the device and then quickly recover to its initial potential value due to the continuous biocatalytic conversion of lactate and O 2 in the biofuel system. The overlay of the potential drop peak is displayed in Figure 3C, where a fast ohmic drop and a subsequent slower voltage drop due to capacitance discharge can be observed. These results demonstrate the ability of BSC to withstand a high current density discharge, reaching up to 10 mA cm -2 . The plot of the calculated power density versus discharge current density, shown in Figure 3D, indicates a maximum power density of 1.7 mW cm -2 at 7.5 mA cm -2. The detailed method of power density calculation is presented in Figure S9. These values are 20 times higher than the power delivered by BFC in continuous discharge mode. Such a high power density indicates the great potential of the BSC to power medical devices, such as insulin pumps or wearable electronic devices, e.g., wireless transmitters. [40] To evaluate the long-time operational stability of the device, a 10 ms 1 mA cm -2 current pulse was applied every 30 s for 1 h; the potential evolution is presented in Figure 3E. The BSC can maintain a stable voltage drop and quickly recovers to its initial voltage after each current pulse over the long-time operation. The calculated relative pulse power density is shown in the inset of Figure 3E, where no significant performance decrease is observed over 120 cycles.

Extended pulsed discharge over 12 h is demonstrated in Figure S10, which shows only minor fluctuations in power and capacitance among thousands of cycles. The performance of the BSC was also evaluated by applying a constant discharge current (0.1 mA cm -2 ), as shown in Figure 3F. Without lactate, the device cannot sustain the discharge and was not able to recharge to OCV above 0.3 V, whereas in the presence of lactate, the BSC can sustain constant discharge for ca. 30 min and can be self-charged to the original voltage, dictated by the fuel concentration. It is worth noting that in contrast to the results demonstrated in Figure 2K, the addition of the 10 mM lactate was able to significantly extend the discharge time due to the bioelectrocatalytic reactions that simultaneously charge the BSC.

To highlight the flexibility and stretchability of this wearable self-charging BSC, its mechanical resiliency towards external strain was tested by implementing resistance and CP tests on the SCs before and after stretching. The resistance measurement during continuous stretching is demonstrated in Figure 3G and Video S1. The resistance profile during measuring is shown in Figure 3H, where the resistance is fluctuating around 205 Ω with a 5% deviation that corresponds to the continuous deformation of the electrode during measurement to adjust to the external strain. The exceptional mechanical stability allows the resistance to recover to its original value after numerous mechanical deformations. The electrochemical behavior of the anodes and cathodes was measured before and after 100 cycles of 180° bending (3 mm diameter, inward and outward) and 20% stretching (along lengthwise) via CV (Figure S11), while monitoring their morphological changes via SEM (Figure S12). The results indicate minimal mechanical damage and changes in performance. Figure 3I shows the stability of the potential profile of BSC discharged at 1 mA cm -2 for 125 stretching cycles. A 10 ms current pulse was applied after every 25 cycles. The power was maintained at over 96% of the maximal value after 125 cycles of 20% stretching. The results demonstrate the robustness of the hybrid device, confirming its potential as a power source for on-body applications. To demonstrate the wearable BSC in real-life scenarios, the printed device was mounted on the arm of a volunteer during the stationary cycling exercise (Figure 4A). The whole process consists of three periods: mounting the BSC and start cycling (a), stop cycling (b), and removing the wearable BSC from the arm of the volunteer (c). Three pulses with the 1 mA cm -2 current were performed every 5 min, and the voltage of the device was recorded (Figure 4B). The OCV of the BSC was around 0.17 V before the sweating and reached 0.48 V after 60 min of cycling.

The OCV dropped to around 0.46 V when the volunteer stopped cycling. After 25 min of resting, the printed BSC was removed from the volunteer's arm. The OCV was observed to rapidly regenerate to 0.48 V due to exposure of the device to additional oxygen, enhancing the reduction reaction on the Pt cathode. Even after removing the BSC from the volunteer's arm for 230 min, the OCV of the BSC was still retained at 0.38 V. The pulse output operated at 1 mA cm -2 current for 10 ms during these different periods is shown in Figure 4C. The power density of the wearable BSC increased from 151 µW cm -2 (Point 1) to 343 µW cm -2 (Point 2), then decreased to 208 µW cm -2 when the volunteer stopped exercising (Point 3) and decreased gradually upon removing the BSC from the skin (Point 4). The power density of the pulses in the whole testing process is shown in Figure S13, demonstrating that the power of the device peaks with the exercise and drops steadily to ~150 µW cm -2 after the exercise stopped. Such performance persisted even after the patch was removed from the body, as the residual stored energy continued to be discharged for another 4 h.

The capability of the BSC to empower commercial electronics is demonstrated in Figure 4D(i) using a simple assembled circuit. Two BSC patches are thus connected in parallel, and the output voltage is boosted via a DC-DC voltage booster. The photo image of a flexible DC-DC voltage booster with laser-cut copper wiring is shown in Figure S14. The output signal from the booster was connected to an LED and was controlled by a switch. To visualize the operation of BSCs and simulation an operation duty cycle of 10%, they were connected to the LED, activating the illumination during a 1 s discharge pulse, followed by a 9 s self-recharge (Figure 4D(ii)). The discharge and self-charge current and voltage profiles are illustrated in Figure 4D(iii); a small current drop is observed during the 1s discharge. Near the turn-on voltage of ca.

1.6 V for the LED, the presence of the lactate allows high current discharge, illuminating the LED in bursts, whereas the absence of lactate cannot provide enough current to surpass the turnon voltage of the LED. A larger current drop is observed in the 5-min extended discharge, as shown in Figure S15. The advantage that the pulsed discharge of the BSC exhibits in such use case involves a high-power pulsed discharge that can sustain the short-term power draw and quickly recovers, whereas the current drops exponentially after the extended discharge, which takes a longer period to recover afterward.

Conclusion

In this work, we demonstrated the first example of an all-printed, stretchable dualfunctional BSC for harvesting and storing energy from metabolic sweat lactate. This hybrid device can rapidly self-charge from autonomous lactate/O2 redox reactions and deliver highpower pulses due to its simultaneous function as both a BFC and an SC. A scalable and low-cost screen-printing technique has been used to fabricate the wearable device, merging these BFC and SC functions on a single footprint. The device was fabricated on top of low-elastic modulus adhesive elastic films for maintaining intimate contact with human skin during rigorous exercises.

The design of the "island-bridge" structure relies on a rigid backbone layer that restrains electrode deformation and stretchable, serpentine-shaped silver interconnection for a controlled strain distribution across the device. Compared to the common lactate BFC, the enhanced capacitance of the BSC is endowed to the synergistic effect of the CNT ink with the electrodeposited PPy conductive polymer on the anode and the high-surface-area porous Pt on the cathode. The resulting wearable bifunctional BSC supports high-power pulsed output while maintaining stable cycling performance in human sweat-level lactate. High power of 0.3 mW cm -2 was achieved when the self-chargeable hybrid device is mounted on the arm of a human subject. When paired with voltage regulation components, commercial electronics -such as LEDs -can be successfully illuminated in pulsed mode by designed BSC, suggesting its considerable potential as a power source for wearable electronics. As a wearable device with enzymatic electrodes, its stability may be affected by environmental factors, such as temperature, pH, salt ions, or surfactants, which will be assessed in the future. Future efforts will further optimize the operational conditions, cell performance, and electronics integration for advancing the capabilities and applications of this novel dual-functional self-charging device.

Formulation of the Stretchable Ag ink and CNT Ink

The preparation of the stretchable Ag and CNT ink is similar to that reported in our previous work [21] . The synthesis of Ag ink comprises the preparation of a polymeric resin consisting of 0.4 g mL -1 of SIS in toluene, then 0.7 g of the resin was mixed with 1.4 g of silver flakes in a dual asymmetric centrifugal mixer (Flacktek Speedmixer, DAC 150.1 KV-K, FlackTek, SC, USA) under a speed of 1800 rpm for 5 min. To enhance the mixing of the ink, Zirconia (YSZ) beads (3 mm, Inframat Advanced Materials, CT, USA) were also added. The CNT ink is synthesized by dispersing 100 mg OH-MWCNT in a solution containing 5 mL of THF and 0.5 mL of terpineol by probe sonication (FS-600 N, Ultrasonic processor, SX Ultrasonics, China) for 5 min.

Thereafter, 100 mg of Tecoflex PU beads were added to the mixture and shaken for 12 h. The formulation of the PU interlayer was followed as previously reported [21] , where 4 g of waterbased Polyurethane resin was mixed with 250 mg of crosslinker before its deposition. The formulation of the PU interlayer ink is formulated by mixing 4 g of water-based polyurethane with 250 mg of crosslinker before its deposition.

CNT Electrode Fabrication

A thin layer of Ecoflex was printed on the adhesive side of a Perme-Roll® Lite film and cured at 65 ℃ for 10 min to form the low elastic modulus substrate. The layers were printed onto screen printing of several layers using an MPM-SPM semi-automatic screen printer (Speedline Technologies, Franklin, MA, USA). A 100 μm-thick stainless-steel printing stencils were designed in AutoCAD (Autodesk, USA) and produced by laser-cut (Metal Etch Services, San Marcos, CA, USA). The printing steps process for the BSC are as follows: A rigid "island-like" structure layer was printed using the dielectric ink firstly onto the modified stretchable film and cured at 80 ℃ for 15 min. Thereafter, to enhance the adhesion, the PU interlayer ink was printed on top of the dielectric layer and was cured at 80 °C for 15 min. Then, the CNT ink was printed on top of the PU layer and cured at room temperature for 8 h followed by 30 min at 80 ℃. Finally, a stretchable conductive serpentine "bridge-like" structure layer was printed using the Ag-SIS ink and cured at 60 ℃ for 15 min.

Electrodeposition of PPy and Pt

The electrodeposition of the PPy and porous Pt were performed in the three-electrode system, with Pt wire as the counter electrode and Ag/AgCl (in saturated KCl solution) as the reference electrode. The PPy deposition protocol followed previous work with minor changes. [41] Shortly, a constant potential of 0.8 V vs. Ag/AgCl was applied to the electrodes immersed into a solution 0.15 M pyrrole, 0.2 M K 2 HPO 4 , and 5 mM KClO 4 for 20 s. A similar technique using commercial Pt electroplating solution was used to deposit the Pt cathode. A high potential of E = -0.95 V vs. Ag/AgCl was set for 15 min to deposit the Pt with high porosity. After the electrodeposition, both electrodes were washed using deionized water 4 times and dried at room temperature.

Enzyme Modification of the Anode

The fabrication of the bioanode was performed by consecutively drop-casting solutions onto the CNT-based electrode surface. The sequence and the formulations of the drop-casting solutions for each electrode (0.16 cm 2 ) were as follows: First, a 5 μL 5 mg mL -1 of OH-MWCNTs dispersed in 0.2 M NQ in 9:1 vol/vol ethanol/acetone solution was cast. Subsequently, a 5 µL aliquot of 40 mg mL -1 LOx in 10 mg mL -1 BSA and 5 µL of 1% glutaraldehyde solution were dropped cast and then covered with 5 µL of 1 wt% chitosan in 0.1 M acetic acid. Each of the above-mentioned steps was followed by a room temperature drying before adding the next layer.

The chitosan layer was allowed to dry overnight.

Assembly of the BSC

A layer of Ecoflex® was used to define the active area and insulate the exposed silver. A PVA-PBS electrolyte gel was prepared by dissolving 1 g of PVA in 5 g of deionized water and crosslinked with an equal weight of 3.3 M KOH solution in a desiccator until 80% weight is lost.

After crosslinking, the hydrogel was repeatedly washed with 0.5 M PBS (pH of 7.4) to remove the excess KOH until reaching a neutral pH. The gel can be stored in PBS before use. The hydrogel is of high flexibility and conformity and was used in on-body experiments for sweat collection, establishing a secure interface between the skin and the electrodes (Figure S16).

Conductive stainless steel thin conductive threads (Adafruit, New York, USA) were used to connect the BSC to the output source using a silver conductive epoxy adhesive (8331, MG chemicals, Canada).

Electrochemical Measurements

Electrochemical studies of the BSC were carried out using μAutolab Type II. Oxygen reduction performance of the cathode was characterized using linear sweep voltammetry (LSV) at 5 mV s -1 in 0.5 M phosphate buffer solution (PBS) (pH 7.4). OCV of the BSCs was measured while quantified lactate solution was added to the 0.5 M PBS (pH 7.4). Polarization curves were obtained using LSV by scanning from OCV to 0 V at 5 mV s -1 . The geometrical area of each compartment, 0.64 cm 2 , was used to calculate the areal power density. The stability of the BSC was analyzed by obtaining the polarization curve of the BSC every 2 h for up to 8 h in the presence of 10 mM lactate at room temperature. The ability of the BSC to deliver power in continuous mode was evaluated under galvanostatic discharge. The BSC was first immersed in 10 mM lactate for 3 min to reach stable OCV and then discharged from OCV to 0 V at 1 mA cm - 2 . The capacitance of the BSC was characterized using cyclic voltammetry (CV) and galvanostatic charge/discharge (GCD) techniques. For single electrode characterization, CV was performed in a three-electrode set-up. For the assembled cell characterization, CV was performed in a two-electrode set-up with the cathode as the working electrode and bioanode as the joint counter/ reference electrode. Chronopotentiometry (CP) was used to evaluate the performance of the BSC in the current pulse mode. For practical evaluation of the device performance, the minimum voltage value (V min ) at the end of the pulse period was chosen. Power value was obtained by multiplying the current pulse and V min .

Characterization of the materials and electrodes

Scanning electron microscopy (FEI Quanta FEG 250) was used to obtain morphological information of the printed electrodes. The element composition of the electrodes was characterized by Energy dispersive X-ray (EDX) mapping analysis.

Characterization of the integrated device

Mechanical tests of the BSC were performed using a motorized linear actuator coupled to a programmable controller (A-LST0250A-E01 Stepper Motor and Controller, Zaber Technologies, BC, Canada). The BSC was stretched repeatedly by 20% at a speed of 0.1 cm s -1 from 0% to 20% reversibly. The resistance change was recorded using a Digital multimeter (34411A, Agilent Technologies, CA, USA). The on-body test was performed by attaching the BSC device facing down to the skin of the arm. A thin film of Perme-Roll® Lite was used to hold and insulate the device onto the skin. Stainless steel thin conductive threads were used to connect the device into output, and silver conductive epoxy was used in the interconnections. All the on-body tests were carried out with consent from the subject volunteers, approved by the Human Research Protections Program at the University of California, San Diego, and in strict compliance with the guidelines of Institutional Review Boards (IRB). The LED pulsed illumination test was conducted by connecting 2 BSC patches in parallel and adding the PBS solution with respective lactate concentrations onto the patch. The patch is connected to a BQ25504 Power Management chip (Texas Instrument, TX, USA) with a complementary circuit, with the output connected to a yellow LED (Sparkfun, CO, USA). The connection is controlled manually. A digital multimeter and a potentiostat are used to simultaneously monitor the current and voltage through/across the LED.

TOC content

An all-printed stretchable epidermal biosupercapacitor that employs novel capacitive electrode material to harvest and store bioenergy from human perspiration in a single wearable device. The device can rapidly self-charge and demonstrates high power in pulsed discharges.

Figure 1 .

 1 Figure 1. Schematic illustration of the epidermal BSC patch. (A) Schematic illustration of the skin-mountable wearable dual-functional (energy harvesting and energy storage) BSC device. (B) Photograph of the stretchable dual-functional BSC device on human skin, scale bar: 2 cm. (C) Schematics of the redox energy generation and capacitive energy storage from sweat lactate by the single dual-functional device. (D)-(F) The behavior of the BSC combining a BFC (left) and SC (center) to form a dual-functional self-charging BSC that supports high-power pulse discharge (right).

  Differences of a 2-fold, 3-fold, and a nearly 5-fold increase in capacitance were observed after the functionalization of Pt alone, PPy alone, and of both Pt and PPy, respectively. The immobilization of LOx leads to a slight decrease in capacitance, demonstrated via EIS and GCD methods (FiguresS6 and S7, respectively) due to partial insulation caused by the enzymatic protein shell and the use of chitosan, which reduces the active surface area and increases the interfacial charge transfer resistance of the electrode. Aiming to test the cyclability of the BSC, 500 CV cycles were performed at a scan rate of 50 mV s -1 . The results, displayed in Figure 2I, indicate only a minor capacitance loss of 4% over time. Figure 2J displays cyclic voltammograms of the BSC at scan rates of 5-50 mV s -1 . A rectangular-shaped response, with slight polarization, is observed, indicating the highly capacitive behavior associated with the high surface area and PPy functionalization. In addition, symmetrical charge-discharge behavior was observed across all the current densities used in GCD, Figure 2K. The capacitance of the BSC decreases upon increasing the current density from 0.1 to 1.0 mA cm -2 , as shown in Figure 2L.

(

  Figure 2F Updated)

Figure 2 .

 2 Figure 2. BSC Bioelectrocatalytic and capacitive behavior characterizations. SEM images of (A) printed pristine CNT electrode, (B) CNT electrode with electrodeposited Pt as the cathode, and (C) CNT electrode with electrodeposited PPy for the anode. Scale bar: (A) and (B) 10 µm; (C) 5 µm. (D) The power density versus voltage plots of the stretchable BSC using different lactate concentrations (0, 2.5, 5, 10, and 15 mM) in 0.5 M PBS (pH 7.4). (E) Polarization curves of the BSC in the presence of 10 mM lactate. (F) Long-term stability of the BSC in the presence of 10 mM lactate over 8 h. Inset: power density at 0.38 V over this period. (G-L) BSC capacitive behavior study. (G) CV (scan rate: 100 mV s -1 ) and (H) galvanostatic charge-discharge curves (current density: 0.1 mA cm -1 ) of the BSC in 2-electrode system with different levels of functionalization. (I) The stability of the BSC is tested with 500 CV cycles at the scan rate of 50 mV s -1 (J) CV curves of the fully functionalized BSC at different scan rates. (K) GCD curves of

Figure 3 .

 3 Figure 3. BSC self-charge and pulsed discharge behavior characterizations. (A) Potential profile of BSC discharge at 1 mA cm -2 current pulse in 0 mM and 10 mM lactate. (B) Potential profile of BSC discharge at 1, 2.5, 5, 7.5, and 10 mA cm -2 over 10 ms (C) Overlay of the BSC

Figure 4 .

 4 Figure 4. BSC On-body behavior. (A) The printed wearable BSC was mounted on the arm of a subject. (B) The voltage monitoring of the wearable BSC: (a) during the on-body constant cycling exercise, (b) after cycling exercise, and (c) unloading the BSC from the skin. (C) The pulse outputs of the wearable BSC (1) during cycling exercise, (2) when the voltage output reaches a stable state, (3) after removing from the body, and (4) after removing the wearable BSC from the skin for 2 h. (D) (i) Circuit schematics demonstrate using two BSC patches to power an LED via a voltage booster. (ii) Photograph of the illuminated LED during and after the pulsed discharge. (iii) The pulsed discharge profile of the BSCs in the absence and presence of 10 mM lactate.

22 4 .

 4 Experimental Section4.1 Chemicals and ReagentsPolystyrene-block-polystyrene (SIS), toluene, 1,4-naphthoquinone (NQ), glutaraldehyde bovine serum albumin (BSA), chitosan, Nafion (20%), L(+)-lactic acid, tetrahydrofuran (THF), terpineol, potassium phosphate dibasic (K 2 HPO 4 ), and Ag flakes (10 μm) were purchased from Sigma-Aldrich. Hydroxyl-functionalized multi-walled carbon nanotubes (OH-MWCNTs) (purity >95%, diameter 10-20 nm, length 10-30 μm) were purchased from Cheap Tubes Inc., USA. LOx was purchased from Toyobo, Japan. Styrene ethylene butylene styrene copolymer (SEBS) was obtained from Kraton, TX, USA. Polyurethane (PU) (Tecoflex SG-80A) was obtained from Lubrizol Life Sciences, OH, USA. Ecoflex 00-30 was purchased from Smooth-On, Inc. PA, USA., and the Perme-roll Lite (L34R10) was obtained from Nitto Denko, Japan. Waterbased polyurethane resin Dispercoll U-42 and Bayhydur 302 crosslinker were purchased from Covestro. Ultra-pure deionized water (18.2 M Ω) was used for preparing all aqueous solutions.

Wearable

  Biosupercapacitor: Harvesting and Storing Energy from Sweat Dr. J. L., L. Y., Dr. X. C., Dr. I. J., C. A. S., Y. L., M. L., Dr. Z. L., Dr. L. W., Dr. A. T., Prof. S. X., Prof. S. C.*, Prof. J. W*

Acknowledgments J. L., L. Y., X. C., and I. J. contributed equally to this work. The authors acknowledge support from the Pack Ambition International Région Auvergne-Rhône-Alpes n° 20 006921 01-91830. X.C. is grateful for a Université Grenoble Alpes Ph.D. scholarship and IDEX for funding X.C.'s travel grant. I.J. acknowledges support from Thai Development and Promotion of Science and Technology Talents Project (DPST). C.S.L. acknowledges support from UC MEXUS-CONACYT fellowship. References [1] J. Wang, M.-F. Lin, S. Park, P. S. Lee, Materials Today 2018, 21, 508.

Supporting Information

Supporting Information is available from the Wiley Online Library or from the author.