Chemiluminescence in Analytical Chemistry
.pdf416 |
Kuroda et al. |
determination of hydrogen peroxide was also reported using ODI in a PO-CL system. In this system, solid-phase detection reactor immobilizing lophine derivatives were introduced and a 10-fmol detection limit for hydrogen peroxide was obtained [93]. Phospholipids separated by preparative HPLC were converted to give hydrogen peroxide by FIA with an IMER, in which phospholipase D and choline oxidase were immobilized, and then determined [94]. Certain amines are known to promote the PO-CL reaction, which was utilized for determination of polyamines in tomatoes [95].
Efforts to find and develop new fluorophores being efficiently chemically excited in the PO-CL reaction were carried out. Pyrimido[5,4-d ]pyrimidines together with several fluorescent compounds were evaluated [96]; 2,6-bis[di-(2- hydroxyethyl)amino]-4,8-dipiperidinopyrimido[5,4-d ]pyrimidine (Dipyridamole) and 2,4,6,8-tetrathiomorpholinopyrimido[5,4-d ]pyrimidine (TMP) gave intense CL, the signals being larger than with any other commercially available fluorescent compound tested (Fig. 16). These pyrimido[5,4-d ]pyrimidines were also applied to a PO-CL photographic assay of hydrogen peroxide and glucose by
Figure 16 Fluorophores with efficient chemical excitation in the PO-CL reaction. TMP, 2,4,6,8-tetrathiomorpholinopyrimido[5,4-d ]pyrimidine; DTDCI, 3,3′-diethylthiadicarbo- cyanine iodide.
Chemiluminescence in Liquid Chromatography |
417 |
using a water-soluble oxamide, 4,4′-oxalyl-bis[(trifluoromethylsulfonyl)imino]- trimethylene-bis(4-methylmorpholinium)trifluoromethanesulfonate (MPTQ) [97]. The highly sensitive detection of near-infrared (near-IR) fluorescent dyes using HPLC with PO-CL detection was examined [98]. These dyes are assumed to be suitable for PO-CL detection owing to their low singlet excitation energy. The detection limits for methylene blue, pyridine 1, oxazine 1 and 3,3′-diethylthiadi- carbocyanine iodide (DTDCI) were 120, 27, 31, and 0.19 fmol on column, respectively. DTDCI was found to be the preferred structure for PO-CL detection and its sensitivity was 250 times that obtained by HPLC with conventional fluorescent detection (Fig. 16).
6.4 HPLC-CL Detection Using Ruthenium Complex
A unique CL reagent, tris(2,2′-bipyridyl)ruthenium(II) [Ru(bpy)32 ] for the postcolumn CL reaction, was applied to HPLC detection. The oxidative-reduction reaction scheme of CL from Ru(bpy)32 is shown in Figure 17. When the production of light following an oxidation of Ru(bpy)32 to Ru(bpy)33 at an electrode surface is measured, this CL reaction is termed electrogenerated chemiluminescence (ECL). The CL intensity is directly proportional to the amount of the reductant, that is, the analyte.
Figure 17 Ru(bpy)32 and its CL reaction.
Table 3 |
Drug Analysis by HPLC with CL Detection Using Ruthenium Complex |
|||||
Drug |
|
Separation column |
Detection |
|||
Antihistamines |
Asahipak ODP-50 C |
18 |
|
ECL |
||
|
|
|
|
|
|
|
|
|
(150 |
4.6 mm id) |
|
|
|
Anticholinergic drugs |
PRP-1 (150 2 mm id) or |
ECL |
||||
|
|
Deltabond octyl silica |
|
|||
|
|
(150 |
2 mm id) |
|
|
|
Erythromycin |
Bioanalytical Unijet C |
18 |
ECL |
|||
|
|
|
|
|
|
|
|
|
(150 |
1 mm id) |
|
|
|
Erythromycin derivative |
Inertsil |
ODS-3 |
|
|
CL |
|
(EM523) |
(150 |
4.6 mm id) |
|
(oxidizing reagent and light |
||
|
|
|
|
|
|
irradiation) |
418
Detection limit (on column) |
Ref. |
0.09–0.21 g/mL |
99 |
(8–16 pmol) |
|
0.1–1 g/mL (0.4–3 pmol) |
100 |
7.4 ng/mL (50 fmol) |
101 |
1 ng/mL plasma |
102 |
10 ng/mL urine |
|
.al et Kuroda
Chemiluminescence in Liquid Chromatography |
419 |
Since the order of increasing CL intensity for alkyl amines reacted with Ru(bpy)32 is tertiary amines secondary amines primary amines, pharmaceutical compounds bearing a tertiary amine function (e.g., antihistamine drugs [99], anticholinergic drugs [100], erythromycin [101], and its derivatives [102]) have been sensitively determined after HPLC separation (Table 3). The method was applied to the detection of D- and L-tryptophan (Trp) after separation by a ligandexchange HPLC [103]. The detection limits for D- and L-Trp were both 0.2 pmol per injection. Oxalate in urine and blood plasma samples has also been determined by a reversed-phase ion-pair HPLC (Fig. 18) [104]. Direct addition of
Figure 18 Chromatogram of oxalate in blood plasma. Peaks: A amino acids; B oxalate. (From Ref. 104.)
420 |
Kuroda et al. |
Figure 19 Derivatization of primary amines with DVS.
Ru(bpy)32 to the mobile phase was investigated and compared with conventional postcolumn Ru(bpy)32 addition. The detection limit using oxalate standards with Ru(bpy)32 in the mobile phase was below 0.1 µM, which was significantly superior to the postcolumn technique. The mobile-phase addition method allowed the instrumentation to be simplified and reduced band broadening caused by postcolumn mixing.
Amino acids labeled with DNS-Cl were determined using the Ru(bpy)32 CL reaction after HPLC separation with a reversed-phase column [104, 105]. DNS derivatives are expected to produce intense CL owing to their secondary and tertiary amino groups. The detection limit for DNS-Glu was 0.1 µM (2 pmol/ injection). Although underivatized amino acids could be detected by Ru(bpy)32 CL, the DNS derivatives showed improved detection limits by three orders of magnitude [105]. An approach to convert primary amines to tertiary amines was also reported [106]. In this method, divinyl sulfone (DVS) was used for a cycloaddition reaction of primary amines (Fig. 19). The DVS derivatives after HPLC separation were sensitively detected (e.g., detection limits for propylamine and 3-aminopentane were 30 and 1 pmol, respectively).
6.5 Other CL Detection Methods for HPLC
Adenine, guanine, and their nucleos(t)ides are known to react with glyoxal derivatives to give chemiluminescent compounds [107–110]. Structures of the chemiluminescent species of both nucleic acid bases are still unknown, but the possible pathway of the derivatization reaction with phenylglyoxal to produce CL is shown in Figure 19. The derivatization products exhibit intense CL in an alkaline medium in the presence of the aprotic polar solvent N,N′-dimethylformamide (DMF). Guanine-containing compounds separated by reversed-phase chromatography were detected with CL and their detection limits range from 4 to 53 pmol (Fig. 20) [110].
Chemiluminescence in Liquid Chromatography |
421 |
Figure 20 Possible pathway of the CL reaction between phenylglyoxal and guanine compound, and chromatogram of guanine compounds. Peaks: 1 GTP; 2 GMP; 3 cGMP; 4 guanosine; 5 deoxyguanosine. (From Ref. 109.)
Methods for determination of thiol drugs (i.e., captopril [21–25], penicillamine [26–28], hydrochlorothiazide [24, 25, 29, 30], and tiopronin [31, 32]) have been developed. These methods are based on CL from a cerium (IV) oxidation system sensitized by adequate fluorophores such as quinine and rhodamine B. By using HPLC-coupled CL-flow-injection analysis method, tiopronin and its metabolite 2-mercaptopropionic acid in human urine were sensitively determined with the detection limits of 0.8 and 1 M, respectively [32].
7. CONCLUSIONS
Research on the application of CL detection in HPLC is progressing rapidly owing to its great sensitivity and the simplicity of instrumentation. These advantages are also well suited for miniaturized separation techniques such as capillary liquid chromatography, capillary electrophoresis, and capillary electrochromatography, which will be further extended. Most of the CL detection systems for HPLC so far reported utilize long-standing CL compounds and reactions, and a few meth-
422 |
Kuroda et al. |
ods have been developed based on new principles. We expect that the efforts will be focused not only on applications but also on the development of new CL reactions and reagents. It is expected that in the near future, CL detection techniques will provide a wider range of applications in the fields of life sciences, environmental science, and other areas.
REFERENCES
1.LJ Kricka, GHG Thorpe. Analyst 108:1274–1296, 1983.
2.PJM Kwakman, UATh Brinkman. Anal Chim Acta 266:175–192, 1992.
3.K Nakashima, K Imai. Molecular Luminescence Spectroscopy, Part 3. New York: Wiley, 1993, pp 1–23.
4.RW Frei, L Michel, W Santi. J Chromatogr 126:665–677, 1976.
5.S Katz, WW Pitt Jr, G Jones Jr. Clin Chem 19:817–820, 1973.
6.S Kobayashi, K Imai. Anal Chem 52:1548–1549, 1980.
7.M Sugiura, S Kanda, K Imai. Biomed Chromatogr 7:149–154, 1993.
8.K Nakasima. Bunseki 7:518–524, 1996.
9.K Hayakawa, N Imaizumi, M Miyazaki. Biomed Chromatogr 5:148–152, 1991.
10.K Imai. Chromatogr Sci 48:359–379, 1990.
11.N Hanaoka. J Chromatogr 503:155–165, 1990.
12.TP Whitehead, GHG Thorpe, TJN Carter, C Croucutt, LJ Kricka. Nature (Lond) 305:158–159, 1983.
13.GHG Thorpe, LJ Kricka, E Gillespie, S Moseley, R Amess, N Baggett, TP Whitehead. Anal Biochem 145:96–100, 1985.
14.GHG Thorpe, LJ Kricka. Methods Enzymol 133:331–353, 1986.
15.M Ii, H Yoshida, Y Aramaki, H Masuya, T Hada, M Terada, M Hatanaka, Y Ichimori. Biochem Biophys Res Commun 193:540–545, 1993.
16.H Hori, T Fujii, A Kubo, N Pan, S Sako, C Tada, T Matsubara. Anal Lett 27: 1109–1122, 1994.
17.N Kuroda, R Shimoda, M Wada, K Nakashima. Anal Chim Acta 403:131–136, 2000.
18.LJ Kricka, X Ji. J Biolumin Chemilumin 10:49–54, 1995.
19.LJ Kricka, M Cooper, X Ji. Anal Biochem 240:11–125, 1996.
20.N Kuroda, K Kawazoe, H Nakano, M Wada, K Nakashima. Luminescence 14: 361–364, 1999.
21.XR Zhang, WRG Baeyens, G Van der Weken, AC Calokerinos, K Nakashima. Anal Chim Acta 303:121–125, 1995.
22.Z Xinrong, WRG Baeyens, G Van der Weken, AC Calokerinos, K Nakashima. J Pharm Biomed Anal 13:425–429, 1995.
23.ZD Zhang, WRG Baeyens, XR Zhang, G Van der Weken. J Pharm Biomed Anal 14:939–945, 1996.
24.J Ouyang, WRG Baeyens, J Delanghe, G Van der Weken, W Vandaele, D De Keukeleire, AM Garcia-Campan˜a. Anal Chim Acta 386:257–264, 1999.
Chemiluminescence in Liquid Chromatography |
423 |
25.J Ouyang, WRG Baeyens, J Delanghe, G Van der Weken, D De Keukeleire, W Vandaele, AM Garcia-Campan˜a, AC Calokerinos. Biomed Chromatogr 12:160– 161, 1998.
26.ZD Zhang, WRG Baeyens, XR Zhang, G Van der Weken. Analyst 121:1569–1572, 1996.
27.ZD Zhang, WRG Baeyens, XR Zhang, G Van der Weken. Biomed Chromatogr 9: 287–288, 1995.
28.ZD Zhang, WRG Baeyens, XR Zhang, Y Zhao, G Van der Weken. Anal Chim Acta 347:325–332, 1997.
29.J Ouyang, WRG Baeyens, J Delanghe, G Van der Weken, AC Calokerinos. Talanta 46:961–968, 1998.
30.J Ouyang, WRG Baeyens, J Delanghe, G Van der Weken, D De Keukeleire, AC Calokerinos. Biomed Chromatogr 12:162–163, 1998.
31.Y Zhao, WRG Baeyens, XR Zhang, AC Calokerinos, K Nakashima, G Van der Weken. Analyst 122:103–106, 1997.
32.Y Zhao, WRG Baeyens, XR Zhang, AC Calokerinos, K Nakashima, G Van der Weken, A Van Overbeke. Chromatographia 44:31–36, 1997.
33.K Ishii, M Yamada. Bunnseki:452–459, 1994.
34.OM Steijger, H Lingeman, UAT Brinkman, JJM Holthuis, AK Smilde, DA Doornbos. J Chromatogr 615:97–110, 1993.
35.T Kawasaki, M Maeda, A Tsuji. J Chromatogr 328:121–126, 1985.
36.H Yuki, Y Azuma, N Maeda, H Kawasaki. Chem Pharm Bull 36:1905–1908, 1988.
37.K Nakashima, K Suetsugu, S Akiyama, K Yoshida. J Chromatogr 530:154–159, 1990.
38.K Nakashima, K Suetsugu, K Yoshida, K Imai, S Akiyama. Anal Sci 7:815–816, 1991.
39.SR Spurlin, MM Cooper. Anal Lett 19:2277–2283, 1986.
40.J Ishida, N Horike, M Yamaguchi. Anal Chim Acta 302:61–676, 1995.
41.J Ishida, N Horike, M Yamaguchi. J Chromatogr B 669:390–396, 1995.
42.JR Zhang, AR Cazers, BS Lutzke, ED Hall. Free Radical Bio Med 18:1–10, 1995.
43.T Miyazawa, S Lertsiri, K Fujimoto, M Oka. J Chromatogr A 667:99–104, 1994.
44.M Maeda, S Shimada, A Tsuji. J Chromatogr 515:329–335, 1990.
45.J Ishida, M Yamaguchi, T Nakahara, M Nakamura. Anal Chim Acta 231:1–6, 1990.
46.J Ishida, S Sonezaki, M Yamaguchi. J Chromatogr 598:203–208, 1992.
47.J Ishida, T Nakahara, M Yamaguchi. Biomed Chromatogr 6:135–140, 1992.
48.J Ishida, S Sonezaki, M Yamaguchi, T Yoshitake. Analyst 117:1719–1724, 1992.
49.J Ishida, S Sonezaki, M Yamaguchi, T Yoshitake. Anal Sci 9:319–322, 1993.
50.J Ishida, T Yakabe, H Nohta, M Yamaguchi. Anal Chim Acta 346:175–181, 1997.
51.M Maeda, A Tsuji. J Chromatogr 352:213–220, 1986.
52.TJ Novak, ML Grayeski. Microchem J 50:151–160, 1994.
53.K Honda, K Miyaguchi, K Imai. Anal Chim Acta 177:103–110, 1985.
54.K Imai, H Nawa, M Tanaka, H Ogata. Analyst 111:209–211, 1986.
55.K Nakashima, K Maki, S Akiyama, WH Wang, Y Tsukamoto, K Imai. Analyst 114:1413–1416, 1989.
56.M Stigbrand, E Ponten, K Irgum. Anal Chem 66:1766–1770, 1994.
57.A Nishitani, Y Tsukamoto, S Kanda, K Imai. Anal Chim Acta 251:247–253, 1991.
424 |
Kuroda et al. |
58.B Mann, ML Grayeski. Biomed Chromatogr 5:47–52, 1991.
59.K Hayakawa, R Kitamura, M Butoh, N Imaizumi, M Miyazaki. Anal Sci 7:573– 577, 1991.
60.K Hayakawa, N Terai, PG Dinning, K Akutsu, Y Iwamoto, R Etoh, T Murahashi. Biomed Chromatogr 10:364–350, 1996.
61.S Kobayashi, J Sekino, K Honda, K Imai. Anal Biochem 112:99–104, 1981.
62.DL Walters, JE James, FB Vest, HT Karnes. Biomed Chromatogr 8:207–211, 1994.
63.S Kobayashi, K Imai. Anal Chem 52:424–427, 1980.
64.K Miyaguchi, K Honda, K Imai. J Chromatogr 303:173–176, 1984.
65.WRG Baeyens, J Bruggeman, B Lin. Chromatographia 27:191–193, 1989.
66.WRG Baeyens, J Bruggeman, C Dewaele, B Lin, K Imai. J Biolumin Chemilumin 5:13–23, 1990.
67.K Miyaguchi, K Honda, K Imai. J Chromatogr 316:501–505, 1984.
68.K Hayakawa, N Imaizumi, H Ishikura, E Minogawa, N Takayama, H Kobayashi, M Miyazaki. J Chromatogr 515:459–466, 1990.
69.A Nishitani, S Kanda, K Imai. Biomed Chromatogr 6:124–127, 1992.
70.K Hayakawa, K Hasegawa, N Imaizumi, OS Wong, M Miyazaki. J Chromatogr 464:343–352, 1989.
71.K Hayakawa, Y Miyoshi, H Kurimoto, Y Matsushima, N Takayama, S Tanaka, M Miyazaki. Biol Pharm Bull 16:817–821, 1993.
72.T Kawasaki, K Imai, T Higuchi, OS Wong. Biomed Chromatogr 4:113–117, 1990.
73.K Nakashima, K Suestsugu, K Yoshida, S Akiyama, S Uzu, K Imai. Biomed Chromatogr 6:149–154, 1992.
74.S Uzu, K Imai, K Nakashima, S Akiyama. Biomed Chromatogr 5:184–185, 1991.
75.S Uzu, K Imai, K Nakashima, S Akiyama. Analyst 116:1353–1357, 1991.
76.H Kouwatli, J Chalom, M Tod, R Farinotti, G Mahuzier. Anal Chim Acta 266: 243–249, 1992.
77.M Tod, M Prevot, J Charon, R Farinotti, G Mahuzier. J Chromatogr 542:295–306, 1991.
78.BW Sandmann, ML Grayeski. J Chromatogr B 653:123–130, 1994.
79.G-L Duan, K Nakashima, N Kuroda, S Akiyama. J Clin Pharm Sci 4:22–28, 1995.
80.K Nakashima, C Umekawa, S Nakatsuji, S Akiyama, RS Givens. Biomed Chromatogr 3:39–42, 1989.
81.K Imai, S Higashidate, A Nishitani, Y Tsukamoto, M Ishibashi, J Shoda, T Osuga. Anal Chim Acta 227:21–27, 1989.
82.H Akiyama, T Toida, T Imanari. Anal Sci 7:807–809, 1991.
83.H Akiyama, S Shidawara, A Mada, H Toyoda, T Toida, T Imanari. J Chromatogr 579:203–207, 1992.
84.S Uzu, K Imai, K Nakashima, S Akiyama. J Pharm Biomed Anal 10:979–984, 1992.
85.Y Hamachi, MN Nakashima, K Nakashima. J Chromatogr B 724:189–194, 1999.
86.K Nakashima, M Nagata, M Takahashi, S Akiyama. Biomed Chromatogr 6:55– 58, 1992.
87.S Yoshida, K Urakami, M Kito, S Takeshima, S Hirose. Anal Chim Acta 239:181– 187, 1990.
88.S Higashidate, K Imai. Analyst 117:1863–1868, 1992.
Chemiluminescence in Liquid Chromatography |
425 |
89.GH Ragab, H Nohta, M Kai, Y Ohkura. Anal Chim Acta 298:431–438, 1994.
90.GH Ragab, H Nohta, M Kai, Y Ohkura, K Zaitsu. J Pharm Biomed Anal 13:645– 650, 1995.
91.K Miyaguchi, K Honda, T Toyo’oka, K Imai. J Chromatogr 352:255–260, 1986.
92.K Nakashima, M Wada, N Kuroda, S Akiyama, K Imai. J Liq Chromatogr 17: 2111–2126, 1994.
93.E Ponten, P Appelblad, M Stigbrand, K Irgum, K Nakashima. Fresenius J Anal Chem 356:84–89, 1996.
94.M Wada, K Nakashima, N Kuroda, S Akiyama, K Imai. J Chromatogr B 678:129– 136, 1996.
95.M Katayama. Meiji Yakka Daigaku Kenkyuu Kiyo 24:43–48, 1994.
96.K Nakashima, K Maki, S Akiyama, K Imai. Biomed Chromatogr 4:105–107, 1990.
97.K Nakashima, S Kawaguchi, RS Givens, S Akiyama. Anal Sci 6:833–836, 1990.
98.K Kimoto, R Gohda, K Murayama, T Santa, T Fukushima, H Homma, K Imai. Biomed Chromatogr 10:189–190, 1996.
99.JA Holeman, ND Danielson. J Chromatogr A 679:277–284, 1994.
100.JA Holeman, ND Danielson. J Chromatogr Sci 33:297–302, 1995.
101.JS Ridlen, DR Skotty, PT Kissinger, TA Nieman. J Chromatogr B 694:393–400, 1997.
102.H Monji, M Yamaguchi, I Aoki, H Ueno. J Chromatogr B 690:305–313, 1997.
103.K Uchikura, M Kirisawa. Anal Sci 7:971–973, 1991.
104.DR Skotty, W-Y Lee, TA Nieman. Anal Chem 68:1530–1535, 1996.
105.W-Y Lee, TA Nieman. J Chromatogr A 659:111–118, 1994.
106.K Uchikura, K Kirisawa, A Sugii. Anal Sci 9:121–123, 1993.
107.N Kuroda, K Nakashima, S Akiyama. Anal Chim Acta 278:275–278, 1993.
108.N Sato, K Shirakawa, K Sugihara, T Kanamori. Anal Sci 13:59–65, 1997.
109.N Kuroda, K Nakashima, S Akiyama, N Sato, N Imi, K Shirakawa, A Uemura. J Biolumin Chemilumin 13:25–19, 1998.
110.M Kai, Y Ohkura, S Yonekura, M Iwasaki. Anal Chim Acta 287:75–81, 1994.