As one of the most important neurochemicals in biological systems, ascorbate plays vital roles in many physiological and pathological processes. In order to understand the roles of ascorbate in the pathological process of tinnitus, this study demonstrates an in vivo method for real time monitoring of the changes of ascorbate level in the cochlear perilymph of guinea pigs during the acute period of tinnitus induced by local microinfusion of salicylate with carbon fiber microelectrodes (CFMEs) modified with multiwalled carbon nanotubes (MWNTs). To accomplish in vivo electrochemical monitoring of ascorbate in the microenvironment of the cochlear perilymph, the MWNT-modified CFME is used as working electrode, a microsized Ag/AgCl is used as reference electrode, and Pt wire is used as counter electrode. Three electrodes are combined together around a capillary to form integrated capillary-electrodes. The integrated capillary-electrode is carefully implanted into the cochlear perilymph of guinea pigs and used both for externally microinfusing of salicylate into the cochlear perilymph and for real time monitoring of the change of ascorbate levels. The in vivo voltammetric method based on the integrated capillary-electrodes possesses a high selectivity and a good linearity for ascorbate determination in the cochlear perilymph of guinea pigs. With such a method, the basal level of cochlear perilymph ascorbate is determined to be 45.0 ± 5.1 μM (n = 6). The microinfusion of 10 mM salicylate (1 μL/min, 5 min) into the cochlear decreases the ascorbate level to 28 ± 10% of the basal level (n = 6) with a statistical significance (P < 0.05), implying that the decrease in ascorbate level in the cochlear may be associated with salicylate-induced tinnitus. This study essentially offers a new method for in vivo monitoring of the cochlear perilymph ascorbate following the salicylate-induced tinnitus and can thus be useful for investigation on chemical essences involved in tinnitus.