Biosurfaces are universally covered with fluid microfilms containing reduced glutathione (GSH) and other antioxidants whose putative roles include the detoxification of ambient ozone (O_3). It is generally believed that O_3 accepts an electron from the thiolate GS^(2-) function [pK_a(GS^-) = 8.8] of GSH to produce thiyl GS^(•-) radicals en route to the disulfide GSSG. Here, we report novel electrospray mass spectrometry experiments showing that sulfonates (GSO_3^-/GSO_3^(2-)), not GSSG, are the exclusive final products on the surface of aqueous GSH microdroplets exposed to dilute O_3(g) for ~1 ms. The higher reactivity of the thiolate GS^(2-) toward O_3(g) over the thiol GS^- is kinetically resolved in this time frame due to slow GS^- acid dissociation. However, our experiments also show   that O_3 will be largely scavenged by the more reactive ascorbate coantioxidant in typical interfacial biofilms. The presence of GSSG and the absence of GSO_3^-/GSO_3^(2-) in extracellular lining fluids are therefore evidence of GSH oxidation by species other than O_3

Colussi, A. J.

Enami, Shinichi

Hoffmann, M. R.

English

Caltech Authors - Main

 1
Supplementary Information for: 
 
Ozone oxidizes glutathione to a sulfonic acid  
Shinichi Enami, M. R. Hoffmann, A. J. Colussi* 
W. M. Keck Laboratories, California Institute of Technology, Pasadena, CA, USA 
 
*Corresponding author: Dr. Agustin J. Colussi, W. M. Keck Laboratories, California 
Institute of Technology, Pasadena, CA 91125 USA;  
E-mail: ajcoluss@caltech.edu 
 2
Supplementary Table S1. Best-fit parameters* for the data of Fig. 4A lower panel. 
 
* Using y = y0 + aexp(-bx) + cx ; y = [I]/[I]0 ;  x = [O3(g)], see reference 1 for details. 
** Correlation coefficient 
 
pH a b c R** γ 
3.5 -4.73 ± 0.31 0.00 ± 0.00 -0.002 0.9648 -0.002 
4.7 0.45 ± 0.15 0.00 ± 0.02 -0.004 0.8427 -0.004 
7.3 0.13 ± 0.01 0.10 ± 0.02 -0.002 0.9157 -0.015 
9.2 0.18 ± 0.00 0.61 ± 0.02 -0.006 0.9902 -0.116 
 
 3
Bulk pH
2 3 4 5 6 7 8 9 10
si
gn
al
 in
te
ns
it
y 
/a
.u
.
0
2
4
6
8
10
12
14
GS2- (m/z  = 152.5)
GS- (m/z = 306)
Supplementary Figure S1  pH dependence on mono- and di-anion signal intensities 
of 0.5 mM GSH. 
 4
[O3(g)]/ppm
0 100 200 300 400 500
I/I
0
0.0
0.2
0.4
0.6
0.8
1.0
GS2-
GS-
  
 
Supplementary Figure S2 Plot of reactants GS- and GS2- signals vs. [O3(g)] at 1.0 
mM GSH with (circle) / without (triangle) excess 1.1 M tert-butanol 
 5
 
 
 
 
[O3(g)] / ppm
0 200 400 600
I/I
0
0.1
1
AH- (1 mM AH2)
AH- (1 mM AH2 + 1mM GSH)
GS- (1mM AH2 + 1mM GSH) 
 
 
Supplementary Figure S3 AH- profiles toward O3(g) in 1.0 mM AH2 at bulk pH = 3.5 (closed 
circle) and AH- and GS- profiles in the mixture of 1.0 mM AH2 + 1.0 mM GSH at bulk pH = 3.5 
(open circle and open square, respectively ). 
 6
Reference 
1. Enami, S., Hoffmann, M.R. & Colussi, A.J. Acidity enhances the formation of a 
persistent ozonide at aqueous ascorbate/ozone gas interfaces. Proc. Natl. Acad. 
Sci. U.S.A. 105, 7365-7369 (2008). 
 
 


Ozone Oxidizes Glutathione to a Sulfonic Acid

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Ozone Oxidizes Glutathione to a Sulfonic Acid

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