Proline-based phosphoramidite reagents for the reductive ligation of S-nitrosothiols

Article abstract of DOI:10.1038/ja.2015.144

S-Nitrosothiols (RSNOs) have many biological implications but are rarely used in organic synthesis. In this work we report the development of proline-based phosphoramidite substrates that can effectively convert RSNOs to proline-based sulfenamides through

Full text of DOI:10.1038/ja.2015.144

The Journal of Antibiotics (2016), 1–6
2016 Japan Antibiotics Research Association All rights reserved 0021-8820/16
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ORIGINAL ARTICLE
Proline-based phosphoramidite reagents
for the reductive ligation of S-nitrosothiols
Chung-Min Park1, Tyler D Biggs1 and Ming Xian
S-Nitrosothiols (RSNOs) have many biological implications but are rarely used in organic synthesis. In this work we report the
development of proline-based phosphoramidite substrates that can effectively convert RSNOs to proline-based sulfenamides
through a reductive ligation process. A unique property of this method is that the phosphine oxide moiety on the ligation
products can be readily removed under acidic conditions. In conjugation with the facile preparation of RSNOs from the
corresponding thiols (RSHs), this method provides a new way to prepare proline-based sulfenamides from simple thiol starting
materials.
The Journal of Antibiotics advance online publication, 13 January 2016; doi:10.1038/ja.2015.144
INTRODUCTION
Small organic thiols (RSHs) can also be easily converted to the
Sulfur-nitrogen (-S-N-) linkages are unique chemical moieties. These corresponding RSNOs under mild nitrosation conditions. Normally,
structures often show interesting bioactivities (Figure 1). For example, RSNOs are unstable species that makes their detection in biological
sulfonylureas are a class of important herbicides.1,2 Sulfanilamides systems very challenging. Moreover, the instability of RSNOs makes
have been used as common drugs for infections.3,4 N-Thiolated these compounds unattractive for synthetic chemists. As a result, the
β-lactams are found to have antibacterial, antifungal and anticancer application of RSNOs in synthesis has been rarely reported. Interest-
effects.5–12 Sulfenamides exhibit antimicrobial activities against various ingly, in our recent work on RSNO bioorthogonal reactions (aiming at
infectious pathogens.13 Proline-based sulfonamides have been the development of novel detection methods for protein RSNO
formation),34–41 we have discovered some unusual reactivity and
properties of RSNOs. In our opinion, RSNOs are powerful synthons
that can be used to introduce S, N and/or O atoms into molecular
structures. Herein, we wish to report a method to prepare proline-
based sulfenamides from readily available RSH substrates via RSNO
intermediates. The novel reactions and synthetic strategies introduced
in this article could be applied for an effective synthesis of bioactive
molecules including a proline moiety.
recognized as potential antiproliferative and anti-infective
agents.14–16 Because of these activities, the preparation of these
molecules, especially the formation of the -S-N- linkages, has become
an active area in organic synthesis. So far, many methods have been
developed for the construction of the S-N bonds, including:
(1) sulfenylation between sulfenyl chlorides and amines,17,18 (2)
amination of thiols with N-halo compounds or amines in the presence
of oxidizing reagents to give unsubstituted and N-substituted
sulfenamides,19 (3) the treatment of disulfides with ammonia or
amines in the presence of silver or mercuric salts in alkaline RESULTS AND DISCUSSION
medium,20,21 (4) sulfenylation of primary and secondary amines with In our previous work, we discovered that RSNOs and triarylpho-
the esters of sulfenic acids,22 (5) derivations of N-chlorothio- sphines (2 equivalents) can rapidly react to generate reactive thioa-
compounds,23,24 (6) the [2,3]-sigmatropic rearrangement of zaylide intermediates in high yields under mild conditions.34
S-allylsulfynimines,25 (7) reductive cleavage of sulfinimidic acids Thioazaylides are potent nucleophilic species. Upon manipulating
in the presence of thiophenols,26 (8) electrolysis of the electrophilic groups attached to the phosphine reagents, thioazay-
2-mercaptobenzothiazole-amine or bisbenzothiazol-2-yldisulfide- lides can be trapped as stable products. For example, as shown in
amine27,28 and (9) the preparation of sulfin- or sulfon-amides by Scheme 1, when an ortho-ester group was attached to the phosphine
the oxidation of sulfenamides in the presence of the oxidants.29,30
reagent (compound 3), the reactive thioazaylide 4 could be trapped via
S-Nitrosothiols (RSNOs) are a group of -S-N- bond containing an intramolecular acyl transfer to afford sulfenamide 5.35–37 This
molecules. The formation of S-nitrosothiols in biological systems is an reductive ligation process provides a way for capturing unstable
important post-translational modification elicited by nitric oxide.31–33 biological RSNO as stable and detectable conjugates. In our opinion,
RSNOs include protein-based adducts (through cysteine residues) and this reaction, in conjunction with easy RSNO formation from RSH,
small molecules (such as S-nitrosoglutathione and S-nitrosocysteine). would be also useful for the synthesis of sulfenamide derivatives from
Department of Chemistry, Washington State University, Pullman, WA, USA
1These authors contributed equally to this work.
Correspondence: Professor M Xian, Department of Chemistry, Washington State University, 468 Fulmer Hall, Pullman, WA 99164, USA.
E-mail: mxian@wsu.edu
Received 9 November 2015; revised 7 December 2015; accepted 13 December 2015

Reductive ligation of RSNOs using proline ester substrates
C-M Park et al
2
Figure 1 Representative molecules having sulfur-nitrogen (-S-N-) linkages.
Scheme 1 Reductive ligations of S-nitrosothiols.
Scheme 2 Reactions between proline-based phosphoramidites and TrSNO.
simple RSH starting materials. However, the use of phosphine suggesting the formation of thioazaylides was fast. For 9 and 10, the
substrates like 3 would lead to products like 5 that contain an reactions stopped at this stage as no desired ligation product was
unnecessary triphenylphosphine oxide moiety. The removal of this obtained even when the reaction time was extended to 24 h. With 11,
bulky group from the final products would require harsh conditions. however, we obtained the desired ligation product 16a. Apparently,
Therefore, seeking phosphine substrates that can undergo the reduc- this can be explained by the factor that phenyl ester is a better leaving
tive ligation while leading to a readily removable phosphine oxide group than the methyl and t-butyl esters. We tested a series of
moiety from the products is desirable. With this idea in mind, we different solvents for this reaction. The best was found to be a mixture
proposed that proline-based phosphoramidites like 6 would be of THF/phosphate buffer (pH 7.4, 20 mM, 3:1) that gave 16a in
suitable substrates. As shown in Scheme 1, the reaction between 84% yield.
RSNO and 6 should still follow the reductive ligation process.
As phosphoramidite 11 was found to be the most reactive substrate
The resultant phosphoramidate moiety can be considered as a for this ligation, it was applied to other RSNO substrates to test the
NH-protecting group and removable under acidic condition. As such, generality of the reaction. A series of primary, secondary and tertiary
proline-linked sulfenamides could be prepared from simple RSH and RSNOs were freshly prepared from RSH and then used in this study
phosphoramidite 6.
without any purification. The results were summarized in Table 1. In
To test this idea, three proline-based phosphoramidites (Scheme 2, all cases, the desired ligation products were obtained. For relatively
9–11) were prepared. These substrates were treated with TrSNO (12a) stable tertiary RSNO substrates, the corresponding ligation products
to explore their reactivity. The selection of TrSNO as the RSNO model were isolated in modest to high yields (entries 1–4). Secondary and
compound was because of its remarkable stability and ease of primary RSNO substrates gave slightly lower yields (entries 5–9).
synthesis. We found the characteristic green color of TrSNO dis- Interestingly, the purification of secondary RSNO products (entries 5
appeared immediately when treated with all three phosphoramidites, and 6) was found to be very difficult because they overlapped with the
The Journal of Antibiotics

Reductive ligation of RSNOs using proline ester substrates
C-M Park et al
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Table 1 Summary of the reactions between 11 and S-nitrosothiols (RSNOs)
aYields detemined by NMR analysis.
phosphine oxide by-product. Nevertheless, the formation of the These results indicated that the reaction indeed proceeded to
products was clearly confirmed by NMR and mass spectroscopy form the thioazaylide intermediate 17. Presumably, the methyl
analysis. These results demonstrated that RSH can be readily converted ester was not reactive enough. Hence, the acyl transfer was slow
to proline-based sulfenamides by this two-step SNO-ligation process. and nonproductive. Instead, the azaylide underwent an intra-
Although phosphoramidite 9 did not show good reactivity to molecular β-elimination on the cysteine substrate to form
convert RSNO to the desired sulfenamides, we did observe some dehydroalanine 18.
unique reactivity of 9. As shown in Scheme 3, the treatment of a
cysteine SNO derivative 12g with gave dehydroalanine 18 sphoryl moiety could be considered as the protecting group of
in a modest yield (37%) together with the disulfide product (46%). proline. We expected it could be removed under acidic
In the proline-based sulfenamide products, the diphenylpho-
9
The Journal of Antibiotics

Reductive ligation of RSNOs using proline ester substrates
C-M Park et al
4
Scheme 3 The reaction between 9 and 12g.
Scheme 4 Deprotection of 16 g.
Scheme 5 The reaction between 11 and HNO.
conditions.42 We then tested the deprotection of 16g (Scheme 4). METHODS
Indeed, a cleavage cocktail of 10% water in TFA provided Instrumentation
1H NMR spectra, 13C NMR and 31P NMR were recorded at 300 MHz (VX 300,
sulfenamide 19 almost quantitatively. This result revealed
that the -S-N- bond on the sulfenamide compounds was quite
stable under acidic conditions. Given the efficiency of this protocol,
it can find applications in making water-soluble proline-based
sulfenamides.
HNO is the one-electron reduced/protonated form of nitric oxide.
It shows distinct physiology and pharmacology from nitric oxide.43
The reductive ligation was also found to be effective for HNO.44
Several specific fluorescent probes for HNO detection have been
developed based on the triarylphosphine template.45–47 We next
wondered whether the proline-based phosphoramidite would work
for HNO and tested the reaction between 11 and HNO (generated
from Angeli’s salt). As shown in Scheme 5, the reaction proved to be
fast and efficient. The desired ligation product 21 was obtained in a
comparable yield (50%) as the one obtained from triarylphosphines,44
suggesting the proline-based phosphoramidite may be useful for the
design of novel HNO sensors.
Varian, Palo Alto, CA, USA) and are reported in p.p.m. on the δ scale relative
1
to residual CHCl3 (δ 7.25 for H and δ 77.0 for 13C). These experiments were
performed at room temperature. Mass spectra were recorded using an
electrospray ionization mass spectrometry (ESI, Thermo Finnigan LCQ
Advantage, San Jose, CA, USA) or MALDI-TOF (matrix-assisted laser
desorption ionization time-of-flight) mass spectrometry. Mass data were
reported in units of m/z for [M+H]+ or [M+Na]+.
Preparation of proline-based phosphoramidites
To a solution of the proline ester HCl salt in CH2Cl2 (total c =0.30 M) was
added freshly distilled 2.5 equivalent of triethylamine followed by PPh2Cl
(1 equivalent) at 0 °C. The mixture was slowly warmed to room temperature
and stirred for 3–4 h. Upon completion (monitored by TLC), the reaction
mixture was filtered to remove the triethylammonium salt. The filtrate was
concentrated and diluted with EtOAc, and washed with satd NaHCO3, water
and brine. The organic layer was dried with anhydrous MgSO4, filtered and
concentrated. The crude product was purified by flash column chromatography
(with preneutralized silica gel by 3% triethylamine in hexanes).
Compound 9: 1H NMR (300 MHz, CDCl3) δ 7.59–7.27 (m, 10H), 4.22 (ddd,
J = 8.8, 6.0, 3.5 Hz, 1H), 3.63 (s, 3H), 3.17–3.06 (m, 1H), 2.88 (m, 1H), 2.22–
1.70 (m, 4H); 31P NMR (122 MHz, CDCl3) δ 49.31; 13C NMR (75 MHz,
CDCl3) δ 176.00, 138.93, 132.88, 132.61, 132.23, 131.98, 128.90, 128.40, 128.33,
128.27, 128.19, 65.28, 64.87, 51.96, 47.64, 47.57, 31.80, 31.71, 25.90; MS (ESI)
m/z calcd for C18H21NO2P [M+H]+ 314.1, found 314.1.
Compound 10: 1H NMR (300 MHz, CDCl3) δ 7.49 (ddd, J = 8.1, 5.4, 1.5 Hz,
2H), 7.39–7.28 (m, 8H), 4.07 (ddd, J = 8.6, 6.6, 3.4 Hz, 1H), 3.17–3.06 (m, 1H),
2.83 (dtd, J = 9.2, 6.8, 2.7 Hz, 1H), 2.07 (dq, J = 12.0, 8.3 Hz, 1H), 1.99–1.82
(m, 2H), 1.76–1.64 (m, 1H), 1.40 (s, 9H). 13C NMR (75 MHz, CDCl3)
δ 174.76, 139.27, 132.93, 132.66, 132.26, 132.01, 128.81, 128.38, 128.29, 128.19,
CONCLUSIONS
In summary, we have developed a reductive ligation of S-nitrosothiols
using N-diphenylphosphine proline ester substrates. This reaction was
found to be effective for all small-molecule S-nitrosothiols (primary,
secondary and tertiary), as well as for HNO. The ligation products
bear a removable phosphine oxide moiety on the proline residue. In
conjugation with the facile preparation of RSNOs from the
corresponding RSHs, this novel method provides a unique way to
prepare proline-based bioactive sulfenamides from simple thiol
starting materials.
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Reductive ligation of RSNOs using proline ester substrates
C-M Park et al
5
128.12, 80.76, 66.37, 65.95, 47.72, 47.65, 31.83, 31.74, 28.24, 25.87. 31P NMR J = 8.5, 6.5, 2.2 Hz, 1H), 3.20 (m, 2H), 2.43 (dd, J = 12.7, 5.9 Hz, 1H),
(122 MHz, CDCl3) δ 49.64.
2.13–2.02 (m, 1H), 1.91 (m, 2H), 1.28 (s, 9H); 31P (122 MHz, CDCl3)
Compound 11: 1H NMR (300 MHz, CDCl3) δ 7.59–7.15 (m, 13H), δ 30.0; 13C NMR (75 MHz, CDCl3) δ 175.98, 132.67, 132.52, 132.39, 132.32,
7.07–6.95 (m, 2H), 4.47 (ddd, J = 8.3, 6.3, 3.8 Hz, 1H), 3.21 (dddd, J = 8.7, 132.19, 129.65, 129.25, 129.13, 129.08, 128.97, 121.40, 62.58, 48.91, 48.35,
7.5, 5.1, 1.0 Hz, 1H), 2.95 (dtd, J =9.4, 7.0, 2.4 Hz, 1H), 2.36–2.14 (m, 2H), 31.49, 28.92, 25.42, 25.33; MS (Maldi) m/z calcd for C21H28N2O2PS [M+H]+
2.09–1.93 (m, 1H), 1.83 (m, 1H); 31P NMR (122 MHz, CDCl3) δ 49.99; 403.1609, found 403.1604.
13C NMR (75 MHz, CDCl3) δ 174.0, 150.9, 138.8, 138.7, 138.6, 132.9, 132.7,
Compound 16g: 1H NMR (300 MHz, CDCl3)
δ 8.97 (s, 1H), 8.30
132.2, 131.9, 129.5, 129.0, 128.5, 128.3, 128.2, 125.9, 121.6, 65.6, 65.1, 47.8, (d, J = 8.3 Hz, 1H), 8.09–7.97 (m, 2H), 7.90–7.67 (m, 4H), 7.62–7.36
47.7, 32.0, 31.9, 25.9; MS (ESI) m/z calcd for C23H23NO2P [M+H]+ 376.1, (m, 9H), 5.02 (ddd, J = 8.3, 6.3, 4.3 Hz, 1H), 4.27–4.02 (m, 1H), 3.74
found 376.2.
(s, 3H), 3.32 (dd, J = 14.8, 6.4 Hz, 1H), 3.04 (m, 3H), 2.35–2.17 (m, 1H),
2.01–1.84 (m, 1H), 1.72 (m, 2H); 31P (122 MHz, CDCl3) δ 30.1; 13C NMR
(75 MHz, CDCl3) δ 176.84, 171.32, 167.73, 133.84, 132.72, 132.50, 132.37,
132.21, 132.07, 131.97, 129.21, 129.10, 128.93, 128.65, 127.91, 61.90, 52.96,
51.28, 48.72, 42.00, 30.95, 25.32; MS (ESI) m/z calcd for C28H30N3NaO5PS
[M+Na]+ 574.3, found 574.3.
Preparation of S-nitrosothiols
The thiol starting material (RSH, 0.2 mmol) was dissolved in 1 ml of MeOH
followed by the addition of 1 N HCl (1 ml) at room temperature. To this
solution was then added freshly prepared 1 N NaNO2 (1 ml) in water in dark
(total c = 0.07 M). The color of the reaction was immediately turned to red
(for primary and secondary RSNO) or green (for tertiary RSNO). The mixture
was stirred for 10–15 min at room temperature. Upon completion (monitored
by TLC), the RSNO product was directly extracted with cold diethyl ether
(1 ml× 3) in dark. The organic layers were collected and dried. The solvent was
removed to provide the RSNO product that was then used for the ligation
reaction without further purification.
Compound 16h: 1H NMR (300 MHz, CDCl3) δ 8.92 (d, J = 7.6 Hz, 1H), 8.44
(s, 1H), 7.75 (tdd, J = 12.0, 8.3, 1.4 Hz, 4H), 7.67–7.38 (m, 6H), 7.01
(d, J = 7.2 Hz, 3H), 6.97–6.85 (m, 3H), 5.38 (td, J = 9.5, 3.2 Hz, 1H), 4.63
(td, J = 8.3, 5.0 Hz, 1H), 4.07 (td, J = 7.0, 5.0 Hz, 1H), 3.67 (s, 3H), 3.42
(m, 1H), 3.27–2.99 (m, 3H), 2.84 (dd, J = 13.9, 8.9 Hz, 1H), 2.49 (dd, J = 14.1,
9.9 Hz, 1H), 2.23–2.10 (m, 2H), 2.08 (s, 3H), 2.02–1.76 (m, 3H); 31P
(122 MHz, CDCl3) δ 30.4; 13C NMR (75 MHz, CDCl3) δ 177.22, 171.75,
171.46, 171.04, 136.73, 132.65, 132.52, 132.33, 132.19, 129.31, 129.20, 129.10,
129.04, 128.93, 128.41, 126.81, 62.41, 54.36, 52.55, 49.93, 43.67, 37.95, 25.70,
23.40; MS (ESI) m/z calcd for C32H37N4NaO6PS [M+Na]+ 659.2, found 659.3.
Compound 16i: 1H NMR (300 MHz, CDCl3) δ 8.21 (d, J = 7.4 Hz, 1H), 7.80
(m, 5H), 7.45 (m, 6H), 7.19 (d, J = 8.1 Hz, 1H), 4.93 (m, 1H), 4.75 (m, 1H),
4.50 (m, 1H), 4.01 (ddd, J = 8.2, 5.9, 2.3 Hz, 2H), 3.75–3.57 (m, 3H), 3.29–3.11
(m, 2H), 2.40–2.24 (m, 1H), 2.10–2.03 (m, 1H), 1.98 (m, 3H), 1.92–1.81
(m, 4H), 1.47–1.29 (m, 3H); 31P (122 MHz, CDCl3) δ 30.4; 13C NMR
(75 MHz, CDCl3) δ 171.22, 170.56, 170.56, 170.08, 132.61, 132.57, 132.54,
132.51, 132.42, 132.37, 132.30, 132.24, 131.76, 131.52, 130.05, 129.84, 129.19,
129.10, 129.03, 128.93, 62.06, 62.02, 53.70, 52.63, 48.47, 48.06, 48.01, 42.04,
31.27, 31.19, 25.26, 25.17, 23.24, 17.97, 17.57; MS (ESI) m/z calcd for
C26H35N4NaO5PS [M+Na]+ 569.2, found 569.3.
General reductive ligation procedure
To the freshly prepared RSNO product was added a solution of 2 equivalent of
11 in 3:1 THF-aqueous buffer (pH 7.4, degassed by bubbling with argon). The
final concentration was ~ 0.1 M. The reaction was monitored by TLC and it was
usually completed within 15–30 min at room temperature. The reaction
mixture was extracted with ethyl acetate. The combined organic layers were
washed with water and brine, dried by anhydrous Na2SO4 and concentrated.
The crude product was purified by flash column chromatography.
Thioazaylide 15: 1H NMR (300 MHz, CDCl3) δ 7.81–7.55 (m, 4H),
7.47–7.17 (m, 23H), 6.94–6.80 (m, 3H), 4.35 (td, J = 8.8, 3.5 Hz, 1H), 3.14
(qd, J = 6.5, 2.8 Hz, 2H), 2.30 (m, 1H), 2.13–1.99 (m, 1H), 1.99–1.78 (m, 2H);
31P (122 MHz, CDCl3) δ 18.3; FT-IR (thin film) 3025.4, 2985.3, 1728.8 (strong,
C =O, carbonyl group), 1601.3, 1450.0, 1372.3, 1268.0, 1247.1, 1070.2 cmÀ1
MS (ESI) m/z calcd for C42H37N2NaO2PS [M+Na]+ 687.2, found 687.1.
Compound 16a: 1H NMR (300 MHz, CDCl3) δ 7.81 (s, 1H), 7.76–7.24
(m, 25H), 3.95 (dt, J =9.2, 5.5 Hz, 1H), 3.01 (p, J =8.1 Hz, 1H), 2.66–2.51
(m, 1H), 1.92 (m, 2H), 1.68–1.61 (m, 1H), 1.34–1.26 (m, 1H); 31P (122 MHz,
CDCl3) δ 30.2; 13C NMR (75 MHz, CDCl3) δ 178.99, 150.71, 145.25, 133.24,
133.12, 132.87, 132.74, 132.01, 131.59, 130.54, 130.04, 129.81, 129.51, 128.59,
128.41, 128.22, 128.15, 127.56, 126.42, 126.00, 121.61, 120.73, 115.57, 63.10,
60.11, 49.03, 32.18, 25.76; MS (ESI) m/z calcd for C36H33N2NaO2PS [M+Na]+
611.2, found 611.1.
Compound 16b: 1H NMR (300 MHz, CDCl3) δ 9.55 (t, 5.6 Hz, 1H, NH)),
9.12 (d, J = 9.5 Hz, 1H, NH)), 8.88 (s, 1H, NH), 8.12–6.82 (m, 20H), 5.18
(d, J = 9.0 Hz, 1H), 4.46–4.33 (m, 2H), 4.20–4.02 (m, 1H), 3.44 (m, 1H), 3.31
(m, 1H), 2.39 (m, 1H), 2.17 (m, 3H), 1.36 (d, J = 10.8 Hz, 6H); 31P (122 MHz,
CDCl3) δ 30.2; 13C NMR (75 MHz, CDCl3) δ 173.13, 169.58, 168.43, 157.53,
150.52, 133.19, 133.06, 133.03, 132.99, 132.53, 132.39, 132.21, 132.06, 131.05,
130.90, 130.49, 129.75, 129.68, 129.62, 129.33, 129.16, 129.01, 128.93, 128.85,
128.61, 128.23, 128.12, 127.42, 126.29, 121.54, 121.42, 119.59, 115.85, 62.20,
60.24, 55.76, 47.78, 43.79, 32.61, 25.91; MS (ESI) m/z calcd for
C36H39N4NaO4PS [M+Na]+ 677.2, found 677.1.
Dehydroalanine (Dha) 18: 1H NMR (300 MHz, CDCl3) δ 8.54 (br, 1H), 7.85-
7.81 (m, 2H), 7.57-7.43 (m, 3H), 6.79 (s, 1H), 5.98 (d, 1H, J = 1.4 Hz), 3.88 (s,
3H); 13C NMR (75 MHz, CDCl3) δ 166.0, 165.0, 134.4, 132.3, 131.2, 129.0,
127.1, 109.1, 53.3; MS (ESI) m/z calcd for C11H12NO3 [M+H]+ 206.1,
found 206.2.
Deprotection of the diphenylphosphoryl group
A sulfenamide product (0.05 mmol) was treated with 1 ml of cold 10% water in
TFA at 0 °C (total c = 0.05 M). The resulted solution was stirred for 30 min at
0 °C. Upon completion, the excess TFA was removed with hexanes as the co-
solvent under vacuum. To the remaining mixture was added cold diethyl ether
to solidify the proline-sulfenamide TFA salt. The solid product was further
washed with cold diethyl ether (2 ml × 5) and dried to provide the final
product.
Compound 19: 1H NMR (300 MHz, CDCl3) δ 10.31 (br-s, 1H), 9.70
(br-s, 1H), 9.09 (s, 1H), 7.57–7.35 (m, 5H), 4.91 (m, 1H), 4.64 (m, 1H),
3.74 (s, 3H), 3.38 (m, 2H), 3.23 (m, 2H), 2.32 (m, 1H), 2.01 (m, 3H); 13C
NMR (75 MHz, CDCl3) δ 172.40, 171.21, 168.61, 132.96, 128.96, 128.70,
127.61, 60.54, 53.21, 51.71, 47.05, 40.52, 29.64, 24.59; MS (Maldi) m/z calcd for
C16H22N3O4S [M+H]+ 352.1331, found 352.1320.
1
Compound 16c: H NMR (400 MHz, CDCl3) δ 8.72 (s, 1H), 7.82 (m, 4H),
The reaction between 11 and HNO
7.58–7.40 (m, 6H), 4.18 (t, J = 7.5 Hz, 1H), 3.19 (dd, J =12.2, 6.0 Hz, 2H),
2.65–2.25 (m, 5H), 2.03 (t, J = 11.8 Hz, 2H), 1.90 (dd, J = 13.6, 10.1 Hz, 5H), To an argon sparged mixture of acetonitrile and water was added 11
1.42 (d, J = 7.3 Hz, 3H), 1.25 (d, J = 8.4 Hz, 3H), 1.00 (d, J = 6.0 Hz, 2.5H), (0.21 mmol), and to this stirring mixture was added freshly prepared Angeli’s
0.93 (d, J = 7.1 Hz, 0.5H); 31P (122 MHz, CDCl3) δ 28.70; 13C NMR salt (0.1 mmol, Na2N2O3). The resulting solution was allowed to stir until the
(101 MHz, CDCl3) δ 211.43, 175.80, 132.34, 132.25, 132.15, 131.96, 131.86, reaction was completed (by TLC, or 20 min). The product 21 (50% yield) was
128.89, 128.79, 128.66, 77.32, 77.00, 76.68, 62.25, 57.80, 52.67, 52.24, 48.20, isolated by extraction and flash column chromatography.
36.73, 34.47, 31.11, 31.05, 30.90, 29.74, 25.64, 25.16, 25.10, 22.28, 22.19; MS
(Maldi) m/z calcd for C27H36N2O3PS [M+H]+ 499.2178, found 499.2201.
Compound 16d: 1H NMR (300 MHz, CDCl3) δ 8.64 (br-s, 1H, NH),
7.92–7.72 (m, 4H), 7.52 (dddd, J = 12.8, 9.3, 7.3, 2.3 Hz, 6H), 4.21 (ddd,
Compound 21: 1H NMR (300 MHz, CDCl3) δ 10.04 (br-s, 1H, NH),
7.96–7.68 (m, 5H), 7.64–7.39 (m, 5H), 5.85 (br-s, 1H, NH), 4.03 (ddd,
J = 8.1, 5.7, 2.3 Hz, 1H), 3.18 (m, 2H), 2.46–2.18 (m, 1H), 2.18–1.99 (m, 1H),
1.89 (ddt, J = 13.5, 8.9, 4.2 Hz, 2H); 31P (122 MHz, CDCl3) δ 28.6; 13C NMR
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Reductive ligation of RSNOs using proline ester substrates
C-M Park et al
6
(75 MHz, CDCl3) δ 176.32, 132.66, 132.62, 132.59, 132.55, 132.44, 132.41,
132.31, 132.28, 131.65, 131.39, 129.92, 129.71, 129.22, 129.13, 129.05, 128.96,
62.12, 62.08, 60.45, 48.07, 48.02, 31.29, 31.21, 25.27, 25.18; MS (ESI) m/z calcd
for C17H19N2NaO2P [M+Na]+ 337.1, found 337.1.
22 Petrov, K. A., Rudnev, G. V. & Sorokin, V. D. Sulphenamides and their derivatives. Russ.
Chem. Rev. 59, 832–843 (1990).
23 Mueller, W. H. & Butler, P. E. β-Chloroalkylsulfenamides. Addition of dimethylamino-
sulfenyl chloride to unsaturated hydrocarbons. J. Org. Chem. 33, 2111–2113 (1968).
24 Kűhle, E. One hundred years of sulfenic acid chemistry IIb. Substitution and cyclization
reactions of sulfenyl halides. Synthesis 1971, 617–638 (1971).
25 Koval, I. V. Progress in the chemistry of sulphilimines. Russ. Chem. Rev. 59,
819–831 (1990).
26 Pel'kis, N. P. & Levchenko, E. S. Interation of areniminosulfinic acid-derivatives with
thiophenols. Zh. Org. Khim. 22, 387–391 (1986).
CONFLICT OF INTEREST
The authors declare no conflict of interest.
27 Torii, S., Tanaka, H. & Ukida, M. Electrosynthesis of hetero-hetero atom bonds. 2. An
efficient preparation of (2-benzothiazolyl)- and thiocarbamoylsulfenamides by
electrolytic cross-coupling reaction of 2-mercaptobenzothiazole, bis(2-benzothiazolyl)
disulfide, and/or bis(dialkylthiocarbamoyl) disulfides with various amines. J. Org. Chem.
43, 3223–3227 (1978).
ACKNOWLEDGEMENTS
This work is supported by the NSF (0844931) and the NIH (R01GM088226).
28 Torii, S., Tanaka, H. & Ukida, M. Electrosynthesis of hetero-hetero atom bonds. 3.
Sodium bromide promoted electrolytic cross-coupling reaction of imides with disulfides.
Convenient synthesis of N-(cyclohexylthio)phthalimide, an important prevulcanization
inhibitor. J. Org. Chem. 44, 1554–1557 (1979).
29 Davis, F. A., Friedman, A. J. & Nadir, U. K. Chemistry of the sulfur-nitrogen bond. 14.
Arenesulfenic acids from N-alkylidenearenesulfinamides (sulfinimines). J. Am. Chem.
Soc. 100, 2844–2852 (1978).
1
2
3
4
Chaleff, R. S. & Mauvais, C. J. Acetolactate synthase is the site of action of two
sulfonylurea herbicides in higher plants. Science 224, 1443–1445 (1984).
Wang, J. G. et al. Structure-activity relationships for a new family of sulfonylurea
herbicides. J. Comput. Aided Mol. Des. 19, 801–820 (2005).
Cooper, L. Z., Madoff, M. A. & Weinstein, L. Absorption and excretion of sulfametho-
midine, a "long-acting" sulfonamide. Antibiot. Chemother. 12, 618–629 (1962).
Shah, S. S., Rivera, G. & Ashfaq, M. Recent advances in medicinal chemistry of
sulfonamides. Rational design as anti-tumoral, anti-bacterial and anti-
inflammatory agents. Mini. Rev. Med. Chem. 13, 70–86 (2013).
Turos, E. et al. N-thiolated bicyclic and monocyclic β-lactams. Tetrahedron 56,
5571–5578 (2000).
O’Driscoll, M. et al. Studies on the antifungal properties of N-thiolated β-lactams.
Bioorg. Med. Chem. 16, 7832–7837 (2008).
30 Haake, M., Gebbing, H.
& Benack, H. A simple method for the oxidation of
sulfenamides to sulfinamides. Synthesis 1979, 97–97 (1979).
31 Zhang, Y. & Hogg, N. S-Nitrosothiols: cellular formation and transport. Free Radical
Biol. Med. 38, 831–838 (2005).
32 Lancaster, J. R. Jr. Protein cysteine thiol nitrosation: maker or marker of reactive
nitrogen species-induced nonerythroid cellular signaling? Nitric Oxide 19,
68–72 (2008).
33 Smith, B. C. & Marletta, M. A. Mechanisms of S-nitrosothiol formation and selectivity in
nitric oxide signaling. Curr. Opin. Chem. Biol. 16, 498–506 (2012).
34 Wang, H. & Xian, M. Fast reductive ligation of S-nitrosothiols. Angew. Chem. Int. Ed.
47, 6598–6601 (2008).
5
6
7
8
Prosen, K. R. et al. The impact of fatty acids on the antibacterial properties of
N-thiolated β-lactams. Bioorg. Med. Chem. Lett. 21, 5293–5295 (2011).
Revell, K. D., Heldreth, B., Long, T. E., Jang, S. & Turos, E. N-thiolated β-lactams:
studies on the mode of action and identification of
a primary cellular target in
35 Zhang, J., Wang, H. & Xian, M. Exploration of the "traceless" reductive ligation of
Staphylococcus aureus. Bioorg. Med. Chem. 15, 2453–2467 (2007).
9
Heldreth, B. et al. N-Thiolated β-lactam antibacterials: effects of the N-organothio
S-nitrosothiols. Org. Lett. 11, 477–480 (2009).
substituent on anti-MRSA activity. Bioorg. Med. Chem. 14, 3775–3784 (2006).
10 Turos, E. et al. N-Thiolated β-lactams: a new family of anti-Bacillus agents. Bioorg.
Med. Chem. Lett. 16, 2084–2090 (2006).
36 Zhang, J. et al. Reductive ligation mediated one-step disulfide formation of
S-nitrosothiols. Org. Lett. 12, 4208–4211 (2010).
37 Zhang, J., Wang, H. & Xian, M. An unexpected Bis-ligation of S-nitrosothiols. J. Am.
11 Turos, E. et al. N-thiolated β-lactams: novel antibacterial agents for methicillin-resistant
Staphylococcus aureus. Bioorg. Med. Chem. Lett. 12, 2229–2231 (2002).
12 Mishra, R. K. et al. N-thiolated 2-oxazolidinones: a new family of antibacterial agents
for methicillin-resistant Staphylococcus aureus and Bacillus anthracis. Bioorg. Med.
Chem. Lett. 16, 2081–2083 (2006).
13 Shang, J. L. et al. Synthesis and evaluation of novel sulfenamides as novel anti
methicillin-resistant Staphylococcus aureus agents. Bioorg. Med. Chem. Lett. 23,
724–727 (2013).
14 Crasto, C. F. et al. Synthesis and activity of analogues of the isoleucyl tRNA synthetase
inhibitor SB-203207. Bioorg. Med. Chem. 11, 2687–2694 (2003).
15 Stefanska, A. L., Cassels, R., Ready, S. J. & Warr, S. R. SB-203207 and SB-203208,
two novel isoleucyl tRNA synthetase inhibitors from a Streptomyces sp. I. Fermentation,
isolation and properties. J. Antibiot. (Tokyo) 53, 357–363 (2000).
16 Huang, S. et al. 1,2,4-Triazolylaminoarylsulfonamide derivatives as antiproliferative
agents, their preparation, pharmaceutical compositions, and use in therapy,
WO2006042215A1 (2006).
17 Koval, I. V. Progress in the chemistry of perhalomethanesulphenyl halides. Russ. Chem.
Rev. 60, 830–849 (1991).
18 Koval, I. V. Sylfenyl chlorides in organic synthesis. Russ. Chem. Rev. 64,
731–751 (1995).
19 Koval, I. V. Thiols as synthons. Russ. Chem. Rev. 62, 769–786 (1993).
20 Davis, F. A. et al. Chemistry of the sulfur-nitrogen bond. VI. Convenient one-step
synthesis of sulfenimines (S-aryl thiooximes). J. Org. Chem. 38, 2809–2813 (1973).
21 Davis, F. A. et al. Chemistry of the sulfur-nitrogen bond. 12. Metal-assisted synthesis of
sulfenamide derivatives from aliphatic and aromatic disulfides. J. Org. Chem. 42,
967–972 (1977).
Chem. Soc. 131, 3854–3855 (2009).
38 Pan, J.
& Xian, M. Disulfide formation via sulfenamides. Chem. Commun. 47,
352–354 (2011).
39 Wang, H., Zhang, J. & Xian, M. Facile formation of dehydroalanine from S-nitrosocys-
teines. J. Am. Chem. Soc. 131, 13238–13239 (2009).
40 Pan, J., Devarie-Baez, N. O. & Xian, M. Facile amide formation via S-nitrosothioacids.
Org. Lett. 13, 1092–1094 (2011).
41 Devarie-Baez, N. O., Zhang, D., Li, S., Whorton, A. R. & Xian, M. Direct methods for
detection of protein S-nitrosylation. Methods 62, 171–176 (2013).
42 Ramage, R., Hopton, D., Parrott, M. J., Kenner, G. W. & Moore, G. A. Phosphinamides:
a new class of amino protecting groups in peptide synthesis. J. Chem. Soc. Perkin
Trans. 1, 1357–1370 (1984).
43 Fukuto, J. M. et al. The physiological chemistry and biological activity of nitroxyl (HNO):
the neglected, misunderstood, and enigmatic nitrogen oxide. Chem. Res. Toxicol. 18,
790–801 (2005).
44 Reisz, J. A., Klorig, E. B., Wright, M. W. & King, S. B. Reductive phosphine-mediated
ligation of nitroxyl (HNO). Org. Lett. 11, 2719–2721 (2009).
45 Reisz, J. A., Zink, C. N. & King, S. B. Rapid and selective nitroxyl (HNO) trapping by
phosphines: kinetics and new aqueous ligations for HNO detection and quantitation. J.
Am. Chem. Soc. 133, 11675–11685 (2011).
46 Miao, Z. et al. A selective phosphine-based fluorescent probe for nitroxyl in living cells.
Bioorg. Med. Chem. Lett. 25, 16–19 (2015).
47 Kawai, K. et al.
A
reductant-resistant and metal-free fluorescent probe
for nitroxyl applicable to living cells. J. Am. Chem. Soc. 135, 12690–12696
(2013).
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