Subscriber access provided by READING UNIV
Communication
Organic Electrochemistry and a Role Reversal: Using Synthesis to Optimize Electrochemical Methods Nai-Hua Yeh, Matthew Medcalf, and Kevin D. Moeller J. Am. Chem. Soc., Just Accepted Manuscript • DOI: 10.1021/jacs.8b02922 • Publication Date (Web): 01 Jun 2018 Downloaded from http://pubs.acs.org on June 1, 2018
Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
Page 1 of 5 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Journal of the American Chemical Society
Organic Electrochemistry and a Role Reversal: Using Synthesis to Optimize Electrochemical Methods Nai-Hua Yeh, Matthew Medcalf, and Kevin D. Moeller* Washington University in Saint Louis, Saint Louis, Missouri 63130, United States
Supporting Information Placeholder ABSTRACT: Diblock copolymers are excellent coatings for microelectrode arrays because they provide a stable surface that can support both synthetic and analytical electrochemistry. However, the surfaces that are optimal for synthetic studies are not the same as the surfaces that are optimal for analytical studies. Hence, no one surface provides an ideal platform for both building and analyzing a molecular library. Fortunately, the synthetic chemistry available on a microelectrode array allows a surface that is ideal for synthesis can be converted into one that is ideal for signaling studies; a scenario that allows for the use of an optimized synthetic and analytical surface on a single microelectrode array.
Scheme 1. The Surface of the Array A growing number of synthetic chemists are turning their attention toward defining the role electrochemistry can play in advancing organic synthesis.1 Fewer chemists are making the connection in the opposite direction and defining the role organic synthesis can play in advancing electrochemical methods. This is a shame because the interplay between organic synthesis and electrochemistry holds significant potential for innovation in both areas. In this paper, we show how Pd(0)- and Cu(I)-catalyzed reactions can be used to take advantage of both a stable hydrophobic surface and a hydrophilic surface on the same microelectrode array. Microelectrode arrays are promising tools for probing interactions between small molecule libraries and biological targets.2 Taking advantage of that opportunity requires the total synthesis of a complex molecular surface on an array so that the individual members of a molecular library can be associated with an individual, spatially addressable electrode or set of electrodes in the array. This is particularly challenging for commercial arrays that have over 12,000 electrodes per square centimeter with those electrodes separated from each other by only 33 microns.3 Central to these efforts is the porous polymer coating on the array (Scheme 1) that is used to attach molecules to the surface of the electrodes. Because the arrays are to be used to evaluate molecules in the library over long periods of time and the analysis of larger libraries will require synthesis of the molecules on the arrays themselves, the surface needs to be stable for long periods of time and chemically inert to a wide variety of chemical reagents. For perspective, at the present Cu(I), Cu(II), Pd(0), Pd(II), acids, bases, Ce(IV), Ru(VII), DDQ, OsO4, Sc(III), H2, TEMPO, and amine radical cations have all been used for conducting siteselective reactions on the surface of an array.4-6
Stability with respect to time and chemistry is not the only requirement for the surface. It must also be compatible with the electrochemical requirements of both the synthesis and signaling experiments. This means it must be porous enough for the chemical reagents to reach the electrodes below, stable with respect to the redox mediator used in signaling, and relatively inert with respect to non-specific biological binding events. Two diblock copolymers pictured in Scheme 1 have proven to be particularly useful.7-12 Both contain a hydrophilic block that is functionalized with a cinnamate ester that can be photochemically crosslinked in order to provide greater stability to the surface. Both polymers also contain a polystyrene hydrophobic block that is functionalized so that groups can be added to the surface. They differ in the nature of that functional group with one polymer containing a bromide11 and the other a borate ester.12 Molecules to be studied on the surface of the array are coupled to the surface by replacing either the bromide or the borate ester in the initial polymer. Unfortunately, neither of the polymers is ideal. The arylbromide-based surface is outstanding in terms of chemical stability and supporting synthesis, but it does not support signaling studies well. The more hydrophilic aryl borated-based surface is far superior in this regard.
ACS Paragon Plus Environment
Journal of the American Chemical Society 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 2 of 5
compatible with many of the synthetic reactions needed to build molecules on an array. So, how does one synthesize and analyze a molecular library on an array if the surface optimized for synthesis and the surface optimized for signaling are not the same? Fortunately, this is a problem that can be solved with organic synthesis. After all, the synthetic community has developed reactions that convert arylbromides into arylborates with the use of a Pd(0)-catalyst,16 and Pd(0)-catalysts can be used site-selectively on an arylbromide coated array.15 The combination of these two precedents suggested that the dilemma outlined above can be solved by using both polymers on the same array. One would simply use the arylbromide polymer for constructing the addressable surface and then convert it into the arylborate surface for the subsequent signaling studies. O
Consider the two CV’s illustrated in Figure 1. Both CV’s were recorded for a 8mM K4Fe(CN)6/K3Fe(CN)6 redox couple in an aqueous 1X PBS electrolyte solution at a sweep rate of 400 mV/sec.3-5 The aqueous buffer solution was used because it is consistent with the conditions needed for monitoring small molecule – protein interactions on an array. The red curve was taken with an array coated with the bromostyrene polymer. Note the very small current measured. Since array-based signaling studies measure a drop in the current associated with the redox mediator,5 the use of the bromide surface has little chance for success. The blue CV was recorded using identical conditions but with an array coated with the borate ester based polymer. In this case, it appears that the more hydrophilic borate ester polymer allows for better swelling, a more permeable coating on the array, and superior diffusion of the redox mediator to the electrodes below. The result is a much higher total current and a much better opportunity for success in subsequent signaling studies.5c While it initially looked like the arylborate surface is the better of the two options,13 it is far too reactive to support synthetic efforts in an acceptable manner. For example, if a Cu(I)-catalyzed reaction is to be run site-selectively on the array, then the array is treated with a Cu(II) precursor and the Cu(I) generated electrochemically specifically where it is needed.14 The arylbromide surface is perfect for such experiments because it does not react with the Cu(II)-precursor. Hence, synthetic tools like “clickreactions” can be used nicely on an arylbromide coated array because the surface only undergoes reactions where the Cu(I) is generated. The same is true for the use of Pd(0) on the arrays. In fact, the arylbromide surface was developed partially to avoid background Pd(II) and acid catalyzed reactions that had plagued the earlier sugar-derived surfaces.15 The borate ester surface shows no such stability. It undergoes reactions under acidic and basic conditions preventing the use of either on the array in a siteselective fashion. In addition, it undergoes background reactions in the presence of a variety of catalysts. On an arylborate ester surface, efforts to conduct Cu(I)-catalyzed click reactions failed because the Cu(II)-precursor utilized itself catalyzed the addition of both acetylenes and azides to the polymer.5d In a similar fashion, both Pd(II) catalyzes additions to the borate ester surface negating the use of Pd(0) on an array coated with this polymer. So while the borate ester surface is excellent for signaling, it is not
O B B
Figure 1. The effect of surface conversion on cyclic voltammetry studies.
O Br
O
O B
allyl acetate PdCl 2(dppf), Ph 3P, TBAB, KOAc, DMF -1.7 V, 4 times (90s) Wholeboard
O
Scheme 2. Pd(0)-catalyzed Aryl Borate Ester Surface Formation For this reason, the reaction in Scheme 2 was explored. In this reaction, the electrodes in an array coated with the arylbromide polymer were used as cathodes to reduce a Pd(II) precursor to Pd(0) at every electrode in the array in the presence of a diborate ester substrate. This was done by setting the potential at each electrode in the array to -1.7 volts relative to a Pt-counter electrode. The reactions are performed as thin film flow reactions with the electrodes in the array serving as the working electrodes and a 0.75 cm2 Pt-plate held at a distance of 650 to 800 mm away from the array with an O-ring serving as the counter electrode.4,5 Tetrabutylammonium bromide was used as the electrolyte for the reaction along with potassium acetate to scavenge the acid generated at the anodic counter electrode. The result of the reaction was to convert the arylbromide surface into an arylborate surface by each electrode. Evidence for the conversion was gathered by examining the CV before and after the conversion. In fact, this is exactly how the image shown in Figure 1 was generated. It would not have been a fair comparison to examine CV’s run on two different arrays because of potential differences in the arrays or the thickness of the surface. So the two CV’s were run on the exact same array as a before and after picture of the reaction shown in Scheme 2. The increase in current observed for the borate ester surface was consistent with earlier studies.5c O
O B B
O Br
O
O B
allyl acetate PdCl2(dppf), Ph3 P, TBAB, KOAc, DMF -1.7 V, 4 times (90s) Wholeboard
O
HO Cu(OAc)2, Bu4 NPF6, DMF +2.4 V, 60 cycles (30s on, 10s off) 4 small squares pattern
O 4
Pyrene
2
ACS Paragon Plus Environment
Page 3 of 5 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Journal of the American Chemical Society Scheme 3. A Chan-Lam Coupling Reaction on the Converted Surface
O
O B B
O
O
O
Br
Further evidence for generation of the borate ester surface was obtained by examining the chemical reactivity of the surface. For example, in Scheme 3 the Pd(0)-catalyzed conversion of the arylbromide surface to the arylborate surface was followed with a subsequent Cu(II)-catalyzed Chan-Lam coupling reaction. In this case, the surface conversion was again conducted at each electrode in the array. The subsequent Chan-Lam reaction conducted at four selected squares of electrode having 840 electrodes (28X30) each.17 For the experiment shown, a Chan Lam reaction was used to put a pyrene group down on the array, and then the success of the reaction was examined with the use of fluorescence microscopy. The image provided shows that the Chan Lam reaction occurred as expected and was confined to only the electrodes selected for the reaction. Since the Chan Lam coupling reaction is not compatible with an arylbromide substrate, it was clear from the image that the surface had been converted to the arylborate. Furthermore, it needs to be noted that to run a Cu(II) reaction selectively on the arrays means treating the array with Cu(I) and then generating the Cu(II) where needed. This is important to note because Cu(I) nicely catalyzes the addition of alcohols to the arylbromide surface.3,5b Hence, the lack of fluorescence from the electrodes not selected for the Chan-Lam reaction in Scheme 3 shows that there was no arylbromide on the surface of those electrodes. The initial conversion arylbromide to arylborate conversion reaction run at those electrodes must have proceeded to completion. As outlined in the supporting information, this is not surprising based on the 360 sec reaction time used for the transformation shown (4 cycles of 90 seconds each). When the time for the reaction was varied, it was found that in all cases the bromide to borate ester conversion was complete in 300 seconds. 18
a) O
O B B
O
O
O
Br
B MeCN:DMF:H2O = 7:2:1 CuSO 4, PPh 3 , Bu4NBr, air -1.7 V, 4 times (90s) Wholeboard
O
HO Cu(OAc)2 , Bu4NPF 6, DMF +2.4 V, 40 cycles (30s on, 10s off) 4 small squares pattern
O 4
b)
Pyrene
B MeCN:DMF:H 2O = 7:2:1 CuSO 4, PPh3 , Bu4 NBr, air -1.7 V, 4 times (90s) Wholeboard
O
OH HO
(PhNH) 2, Bu 4NPF 6 , MeOH, pyridine +2.4 V, 20 cycles (30s on, 10s off) 4 small squares pattern
O B
Pyrene
O 3
Scheme 4. Cu(I)-catalyzed Arylborate Ester Surface Formation For an array based molecular library it is advantageous if the surface can be tuned in order to minimize non-specific binding events between the polymer and a targeted protein. To do so requires both an irreversible reaction to place the molecules to be studied onto the array proximal to the electrodes13 and then a reversible reaction to subsequently tune the surface in response to the various proteins that might be studied.12 Furthermore, the presence of trace amounts of Pd can interfere with biological studies on the array. What the two reactions in Scheme 4 illustrate is that first a Cu(I)-catalyst can be used to convert the arylbromide surface to the arylborate surface and in so doing the use of Pd can be avoided altogether. Second, the two reactions show that both an irreversible Chan-Lam coupling13 and a reversible alcohol exchange reaction12 can be conducted site-selectively on the new surface of the array in a fashion identical to the chemistry conducted on arrays coated directly with the borate ester copolymer. In the experiments shown above, the whole array was used for the reaction to convert the arylbromide to the arylborate. This was done because for most needs a molecular library will be built on the array and then the surface converted to the borate ester. However, a scenario where one might want to build part of a library, convert the surface located by that portion of the library to a borate ester, gather initial binding data on the first part of the library, and then use that data to guide the synthesis of the remainder of the library. For such a scenario, the arylbromide to arylborate surface conversion needs to be done in a site-selective manner on the array. This is not a problem because Pd(0)- and Cu(I)catalyzed reactions can be readily confined on the arylbromide surface,15 and this proved true for the arylborylation as well.18 In conclusion, we have found that the development of new synthetic methodology on a microelectrode array allows for the use of both an optimal array surface for synthesis and an optimal surface for signaling on the same array even though those two surfaces are not the same.
ASSOCIATED CONTENT Supporting Information Sample experimental procedures for the site-selective reactions are included along with the protocol for conduction cyclic voltammetry studies on the arrays. This material is available free of
charge via the Internet at http://pubs.acs.org.”
3
ACS Paragon Plus Environment
Journal of the American Chemical Society 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
AUTHOR INFORMATION Corresponding Author
[email protected] Notes The authors declare no competing financial interests.
ACKNOWLEDGMENT We thank the National Science Foundation (CBET 1262176) for their generous support of our work.
REFERENCES (1) Yan, M.; Kawamata, Y.; Baran, P. S. Chem. Rev. 2017, 117,1323013319. (2) (a) Sullivan, M. G.; Utomo, H.; Fagan, P. J.; Ward, M. D. Anal. Chem., 1999, 71, 4369-4375. (b) Zhao, H.; John, R. Anal. Chim. Acta 2000, 421, 175-187. (c) Hintsche, R.; Albers, J.; Bernt, H.; Eder, A. Electroanalysis 2000, 12, 660-665. (d) Gardner, R. D.; Zhou, A.; Zufelt, N. A. Sensors and Actuators B 2009, 136, 177-185. (e) Beyer, M.; Nesterov, A.; Block, I.; König, K.; Felgenhauer, T.; Fernandez, S.; Leibe, K.; Torralba, G.; Hausmann, M.; Trunk, U.; Lindenstruth, V.; Bischoff, F. R.; Stadler, V.; Breitling, F. Science 2007, 318, 1888. (f) Devaraj, N. K.; Dinolfo, P. H.; Chidsey, C. E. D.; Collman, J. P. J. Am. Chem. Soc. 2006, 128, 17941795. (g) Wassum, K. M.; Tolosa, V. M.; Wang, J.; Walker, E.; Monbouquette, H. G.; Maidment, N. T. Sensors 2008, 8, 5023-5036. (h) Kerkoff, H. G.; Zhang, X.; Mailly, F.; Nouet, P.; Liu, H.; Richardson, A. VLSI Design 2008, 2008 (Article ID 437879), 9. (i) Zhang, Y.; Wang, H.; Nie, J.; Zhang, Y.; Shen, G.; Yu, R. Biosensors and Bioelectronics 2009, 25, 34-40. (j) Maurer, K.; Yazvenko, N.; Wilmoth, J.; Cooper, J.; Lyon, W.; Danley, D. Sensors 2010, 10, 7371-7385. (k) Li, X.; Tian, Y.; Xia, P.; Luo, Y.; Rui, Q. Anal. Chem. 2009, 81, 8249-8255. (l) Chan, E. W. L.; Yousaf, M. N. ChemPhysChem 2007, 8, 1469-1472. (m) Chan, E. W. L.; Yousaf, M. N. J. Am. Chem. Soc. 2006, 128, 15542-15546. (n) Roth, K. M; Peyvan, K.; Schwarzkopf, K. R.; Ghindilis, A. Electroanalysis 2006, 18, 1982-1988. (o) Arya, S. K.; Chornokur, G.; Venugopal, M.; Bhansali, S. Biosensors and Bioelectronics 2010, 25, 2296-2301. (3) For a description of the chips used here see: Dill, K.; Montgomery, D. D.; Wang, W.; Tsai, J. C. Anal. Chim. Acta 2001, 444, 69-78. 12K slide: diameter = 44 µm; Distance between the Pt-electrodes (square cells) = 33 µm. For a detailed discussion of how the array reactions are run see the supporting information for Bartels, J.; Lu, P.; Maurer, K.; Walker, A. V.; Moeller, K. D. Langmuir 2011, 27, 11199-11205 and references 4 and 5b below. (4) For an instructional review see: Graaf, M. D.; Moeller, K. D. Langmuir 2015, 31, 7697-7706. (5) For examples see: (a) Stuart, M.; Maurer, K.; Moeller, K. D. Bioconjugate Chem. 2008, 19, 1514-1517. (b) Stuart-Fellet, M.; Bartels, J. L.; Bi, B.; Moeller, K. D. J. Am. Chem. Soc. 2012, 134, 16891-16898. (c) Uppal, S.; Graaf, M. D.; Moeller, K. D. Biosensors 2014, 4, 318-328. (d) Graaf, M. D.; Marquez, B. V.; Yeh, N. –H.; Lapi, S. E.; Moeller, K. D. ACS Chem. Biol. 2016, 11, 2829-2837. (6) (a) Bi, B.; Huang, R. Y. –C., Maurer, K.; Chen, C.; Moeller, K. D. J. Org. Chem. 2011, 76, 9053-9059. (b) Bi, B.; Maurer, K.; Moeller, K. D. J. Am. Chem. Soc. 2010, 132, 17405-17407. (7) For examples using agarose see: (a) Bi, B.; Maurer, K.; Moeller, K. D. Angew. Chem. Int. Ed. 2009, 48, 5872-5874. (b) Bartels, J. L.; Lu, P.; Walker, A.; Maurer, K.; Moeller, K. D. Chem. Commun. 2009, 55735575. (8) For examples using sucrose see: Maurer, K.; McShea, A.; Strathmann, M.; Dill, K. J. Combi. Chem. 2005, 7, 637-640. (9) For an example of agarose instability see: Kesselring, D.; Maurer, K.; Moeller, K. D. J. Am. Chem. Soc. 2008, 130, 11290-11291. (10) For the basic polymer used: (a) Cambre, J. N.; Roy, D.; Gondi, S. R.; Sumerlin, B. S. J. Am. Chem. Soc. 2007, 129, 10348-10349. (b) Pennington, T. E.; Kardiman, C.; Hutton, C. A. Tetrahedron Lett. 2004, 45, 6657-6660.
Page 4 of 5
(11) For the phenylbromide based suface used on an array: Hu, L.; Bartels, J. L.; Bartels, J. W.; Maurer, K.; Moeller, K. D. J. Am. Chem. Soc. 2009, 131, 16638-16639. (12) For the borate ester derived copolymer used on an array: Hu, L.; Graff, M. D.; Moeller, K. D. J. Electrochem. Soc. 2013, 160, G3020G3029. (13) Graaf, M. D.; Moeller, K. D. J. Org. Chem. 2016, 81, 1527-1534. (14) Bartels, J.; Lu, P.; Maurer, K.; Walker, A. V.; Moeller, K. D. Langmuir 2011, 27, 11199-11205. (15) Hu, L.; Stuart, M.; Tian, J.; Maurer, K.; Moeller, K. D. J. Am. Chem. Soc. 2010, 132, 16610-16616. (16) (a) Ishiyama, T.; Murata, M.; Miyaura, N. J. Org. Chem. 1995, 60, 7508-7510. (b) Kleeberg, C.; Dang, L.; Lin, Z.; Marder, T. B. Angew. Chem. Int. Ed. 2009, 48, 5350-5354. (17) Since the electrodes in the arrays are individually addressable any pattern can be used for a reaction. In this case, the number of electrodes used was arbitrary and selected simply because the pattern was easy to see both under a microscope and if confined well using ones “naked eye”. (18) Details are included in the supporting information.
4
ACS Paragon Plus Environment
Page 5 of 5 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Journal of the American Chemical Society
TOC Graphic
5
ACS Paragon Plus Environment
