Cyclic voltammetry and chronoamperometry at micropipet electrodes were applied to study the phase transfer of polypeptide protamine facilitated by complexation with charged ionophore dinonylnaphthalenesulfonate (DNNS) at polarized water/1,2-dichloroethane (DCE) interfaces, i.e., sDNNS(-) (DCE) + protamine(n+) (aq) right harpoon over left harpoon protamine-DNNS complex (DCE). Well-defined current responses based on the selective protamine transfer were obtained reproducibly even in the presence of 0.12 M NaCl. The selective and reproducible responses make this voltammetric/amperometric approach an attractive alternative to the traditional potentiometric counterpart based on mixed potential responses, for which both protamine and Na(+) need to be transferred simultaneously. Using both organic- and water-filled micropipet electrodes, the reaction mechanism was studied under different mass-transfer conditions controlled by diffusion of protamine, DNNS, and the complex in the outer solution of the pipets. Both charge number of transferred protamine, n, and complexation stoichiometry, s, were determined to be approximately 20 by chronoamperometry. With these parameters, the electrochemically irreversible voltammograms were analyzed by assuming a one-step transfer model to obtain experimental transfer coefficients, which represent apparent dependence of the transfer rate on the interfacial potential. The analysis showed that the transfer coefficients are much larger or smaller than a normal value of approximately 0.5 and strongly depend on the diffusion-limiting species, i.e., 0.088 +/- 0.005, 0.89 +/- 0.01, and 0.065 +/- 0.008 for protamine, DNNS, and the complex, respectively. The apparently anomalous transfer coefficients were explained consistently by a phenomenological model based on adsorption and transfer processes.

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Heparin, a highly negatively charged polysaccharide, which has been used widely as an anticoagulant and antithrombotic, was detected by ion-transfer voltammetry at the interface between 1,2-dichloroethane and an aqueous buffer solution or undiluted blood plasma. Quaternary ammoniums with different numbers of methyl and long alkyl groups were examined as positively charged heparin ionophores using pipet electrodes filled with the organic electrolyte solutions of their tetrakis(4-chlorophenyl)borate salts. It was shown that octadecyltrimethylammonium most selectively facilitates interfacial heparin adsorption without interference from potential-dependent ionophore transfer into the aqueous phase. Water-filled pipet electrodes were also used to study the stoichiometry of the interfacial complex between a heparin molecule and multiple ionophore molecules, which is discussed as a counterion condensation effect. Stripping voltammetry based on facilitated heparin adsorption and desorption gives a detection limit of 0.012 unit/mL in 0.12 M NaCl buffered at pH 7.2, which is 1 order of magnitude lower than therapeutic heparin concentrations (>0.2 unit/mL) and is comparable to a detection limit of the most sensitive heparin sensor reported so far. The biomedical utility of ion-transfer voltammetry was demonstrated for the first time in an undiluted blood sample. Despite interferences by Na+, Cl-, and plasma proteins such as serum albumin, a detection limit of 0.13 unit/mL was obtained in sheep blood plasma with the stripping method.