Biomedical Engineering Reference
In-Depth Information
stocks of 10 mM were made by dissolving fresh crystals of Na
2
S in the same analytical
grade water in the following procedure. The buffer solution in a 20 ml Pyrex syringe
was vigorously sparged with argon for at least 10 min to achieve anoxia. Liquid from
the syringe was then used to dissolve the crystals in a conical Pyrex centrifuge tube
fi lled with a continuous stream of argon, pH was adjusted to 7 with dilute HCl, and
the solution was immediately drawn back into the syringe free of bubbles and sealed
with a rubber serum stopper. Aliquots of the anoxic Na
2
S stock were obtained with
a gas-tight syringe (Hamilton, Reno, NV) through the stopper and injected into the
respirometer. Dilute H
2
S stocks of 0.1 mM were made by injecting aliquots of the con-
centrated stock through the sealing stopper of a second syringe containing anoxic buff-
ered water. Stock solutions were calibrated with the standard 2,2
-dipyridyl disulfi de
(2-PDS) assay [43]. In this assay, samples of the H
2
S stock were dissolved into an
excess of the 2-PDS reagent so that all H
2
S reacted to form stoichiometric amounts of
the product 2-thiopyridone; one H
2
S produces two 2-thiopyridone. Concentration of
2-thiopyridone was determined by dividing the optical density at 343 nm of the assay
solution in a 1 cm pathlength cuvette by the extinction coeffi cient of 8.08 mM
1
cm
1
[44]. A regression of the expected H
2
S concentration vs the measured 2-thiopyridone
concentration yielded a slope that was typically within
2% of unity. H
2
S stock solu-
tions were also prepared by equilibrating anoxic buffer with H
2
S gas (Mattheson),
yielding concentrations near 140 mM H
2
S. These stocks were used for very small vol-
ume injections with negligible effects on pH.
8.4.2 Chemical sources of H
2
S
In studies in which NaHS is used as the sulfi de source, effective concentrations are
typically higher than with other sulfi de sources [20]. Stock solutions prepared with
yellow NaHS fl akes are pale yellow in color compared to the clear solutions prepared
with Na
2
S or equilibrated with H
2
S gas, suggesting the presence of elemental sulfur in
NaHS solutions. The PHSS consistently produced a twofold higher signal in response
to Na
2
S injections compared to injections of the same concentration of NaHS [41],
indicating that, although the yellow NaHS solution produced a linear signal, approxi-
mately one half of the sulfur is present in a form not able to diffuse through the H
2
S-
permeable membrane to chemically react with the ferricyanide electrolyte. Spectral
features of the anoxic stock solutions, specifi cally a strong 380 nm absorbance band
exhibited by the NaHS solution compared to little 380 nm absorption by the Na
2
S solu-
tion, suggested the presence of scattering components such as polysulfi des or elemen-
tal sulfur in NaHS solutions.
8.5 CHARACTERIZATION OF POLAROGRAPHIC
H
2
S SENSORS
PHSS performance characteristics are described below and compiled in Table 8.1.
PHSS performance and calibration were evaluated in the temperature-controlled
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