HPLC is utilized extensively as a means of detecting and determining trace components. In particular, it is effective in the case of involatile trace components in complex matrix such as biological substances. Labeling objective substances for analysis with labeling reagents appropriate for detection methods, has been performed in order to obtain higher sensibility and selectivity. A great number of labeling reagents have been reported for this purpose.
For example, NBD- and DBD-derivatives are excellent fluorescent labeling reagents with strong and long wave-length fluorescence resulting from their benzoxadiazole skeleton and they are used for analysis of various biological substances.^1,2) N-(4-Aminobutyl)-N-ethylisoluminol has been reported to be usable as a chemiluminescence labeling reagent.³⁾

For electrochemical detectors (ECD), ferrocene derivatives and NPCA⁴⁾ have been reported. Ferrocenes have been utilized as the most suitable labeling reagents for ECD because they respond to the application of relatively low voltages, such as +0.5 V, and they are excellent redox system of oxidation and reduction. NPCA has been developed by the research group of University of Shizuoka recently. Interaction of NPCA with various primary amines under mild conditions, followed by removal of the THP group affords the corresponding derivatives which have 6-hydroxychroman skeleton. These derivatives exhibit intense electrochemical activity based on 6-hydroxychroman skeleton and can be separated efficiently by HPLC on an ODS column. The simultaneous determination of neuroactive monoamines and their metabolites is possible in this HPLC-EC method, and it is expected to be applied in neurochemical research.

UV Detection

Fluorescence Detection

Chemiluminescence Detection

Electrochemical Detection

UV Detection

for Carboxyl Groups

A5507	B0949	B0633
A5505	M0815	A5501
A5502	A5503	A5504
A5506	A5508

for Amino Groups

N0713	N0714	I0125
I0190	A5514	A5515
A5512	A5511	D1653
M0721	N0176	F0197
B0105	D1382	I0189
A5522	A5524	D2259
A5523	I0015	I0131
A5513	P1662	S0004

for Hydroxyl Groups

N0713	N0714	A5511
M0721	N0176	B0105
I0131

for Carbonyl Groups

D0845	A5531	A5533
A5532

Fluorescence Detection

for Carboxyl Groups

A5561	A5563	A5560
A5562	A5553	A5570
A0979	B1926	A5502
A5576	A5551	A5571
A5574	A5555	A5573
A5554

for Amino Groups

A1657	N0713	A1658
N0714	A5565	A5566
A5568	A5577	A5569
A5578	A5564	A5567
A5594	M0722	A5579
D3178	F0197	D0005
D0656	A5558	A5595
A5575	F0192	F0026
A5592	A5572	A5593
P0280	S0503

for Hydroxyl Groups

A1657	N0713	A1658
N0714	A5565	A5566
A5564	A5567	A0411
A5579	D3178	D0005
D0656	A5558	A5572

for Carbonyl Groups

A5581	A0267	A5552
A5571	A5556	A5573
A5557

for Mercapto Groups

A5597	A5596	A5595
P1214	A5591	A5592
A5593

Chemiluminescence Detection

D2339

A5304

Electrochemical Detection

H0941	F0406	F0280
S0820	S0599