the latter required the complexation of metal ions to an electroactive ligand (dithiocarbamate). Once the electroinactive complex was formed, the con sumption of the reagent was monitored electrochemically. Although these re ports indicate the feasibility of indirect measurements, the sensitivity of the measurement must be enhanced through careful selection of the electrophore and further optimization of the electrochemical dynamic reserve. Ewing and co-workers (28-30) dem onstrated that extremely sensitive am perometric measurements can be made in CZE by electrically decoupling an amperometric detector from the electrophoretic power supply. Basically, a small break in the separation capillary several centimeters before the detector serves as a conductive joint, which re moves the current from the separation without disturbing the flow of buffer along the capillary. A carbon fiber (ra dius 5-10 Mm, length 100-500 Mm) is inserted into the end of the column and the reference electrode is placed in buffer at the end of the capillary, thus forming an amperometric detector with a volume of only a few tens of picoliters. The apparatus is housed in a Faraday cage to minimize noise.
ACS
Because nanomolar detection limits were found for indoles and catechol amines (28), indirect electrochemical detection uses a similar experimental paradigm. Olefirowicz and Ewing (35) demonstrated the feasibility of indirect amperometric detection using 26 -μπιi.d. capillaries and 5-jum-diameter carbon fiber working electrodes. Dihydroxybenzylamine (DHBA) was used as the cationic electrophore, which pro vided a stable background at concen trations as low as 10 μΜ in the presence of 25 mM 2-morpholinoethanesulfonic acid (MES) (Figure 5). Several amino acids and peptides were detected in this manner with 500-attomole detec tion limits. Direct amperometric detec tion of catechols could also be per formed simultaneously, allowing both electroactive and electroinactive con stituents of the sample to be monitored in the same run. Alternatively, an ion-selective elec trode (ISE) of micrometer dimensions may serve as a sensitive detector ideal ly suited for indirect detection. Potentiometric measurements of many cat ions have been performed on column in capillary HPLC by placing the micro meter-wide tip of the ion-selective microelectrode directly into the end of
ΟΗΒΑ
Time (min)
Figure 5. Electropherogram of amino acids with indirect detection. Buffer: 0.1 mM DHBA-0.025 M MES (pH 5.50)10% (v/v) ethanol; 72.1-cm-long, 26-μΓπ i.d. sep aration capillary; 0.6-cm detection capillary; in jection by electromigration, 3 s at 15 kV; separa tion voltage, 15 kV; electrode potential, 0.7 V versus SSCE. Peaks: A, Lys; Β, Arg; C, His; S, system peak. (Adapted with permission from Ref erence 35.)
the separation capillary (36). A potassi um-selective microelectrode (5-μπι tip
Atitech Capillary Columns and Accessories
i-3 CD
y/ Quality y/ Value ·>/ Service
5 Bulletin #183
η
FOR OVER 50 PAGES OF COLUMNS, ACCESSORIES AND APPLICATIONS — REQUEST BULLETIN #183
y j lltech manufactures a whole family of capillary columns. Designed for both general and specialty separations, our columns are reasonably priced and meet the most demanding specifications. For Quality, Value, and Service — call Alltech!
Alltech
Alltech Associates, Inc. 2051 Waukegan Road Deerfield, IL 60015 Phone: 1-800-255-8324 · Fax: 708-948-1078
CIRCLE 3 ON READER SERVICE CARD ANALYTICAL CHEMISTRY, VOL. 63, NO. 5, MARCH 1, 1991 • 281 A
