Fluorescence detection, in HPLC

The excitation and emission wavelengths used for fluorescence detection in HPLC analysis are 350 and 450 nm for isoxanthopterin and 340 and 450 nm for 2,4,7-trioxopteridine, respectively." The absorption spectra of riboflavin present... 

Instrumentation for fluorescence spectroscopy has been reviewed [8]. For standards in fluorescence spectroscopy, see Miller [138]. Fluorescence detection in HPLC has recently been reviewed [137], Phosphorescence detection of polymer/additive extracts is not being practised. 

Captamine hydrochloride EINECS 236-221-6 MEDA NSC-45463 Thiofluor , Nucleophile used in preparation of o-phthaldehyde reagent for fluorescence detection in HPLC. Metal complexing agent and depigmentor. Crystals mp = 158-160°. Schering-Plough Healthcare Products. 

The limits of lifetime detection and resolution in on-the-flight fluorescence lifetime detection in hplc were evaluated for simple, binary systems of polycycHc hydrocarbons (70). Peak homogeneity owing to coelution was clearly indicated for two compounds having fluorescence lifetime ratios as small as 1.2 and the individual peaks could be recovered using predeterrnined lifetimes of the compounds. Limits of lifetime detection were deterrnined to be 6 and 0.3 pmol for benzo[b]fluoranthene and benzo[k]fluoranthene, respectively. 

In LIF detection systems, excitation power may be increased up to six orders of magnitude compared to CF detection. Most LC-LIF detection concerns under-ivatised polynuclear aromatic hydrocarbons (PAHs) and fluorescing dyes (e.g. polymethines). Because only a limited number of analytes possess native fluorescence, derivatisation of the analyte before detection is normally required in trace analysis of organic solutes by means of LIF detection. LIF detection in HPLC was reviewed... 

Detectors. Fluorescence and UV detectors are used in the HPLC analysis. The high sensitivity and specificity of fluorescence detection in tocopherols and tocotrienols make the fluorescence detector the first choice. The fluorescence detector is ten times more sensitive and has less background noise than the UV detector. Electrochemical detectors are also used in the analysis of tocopherols and tocotrienols (Murphy and Kehrer, 1987 Sanchez-Perez et al., 2000). As a high-polarity mobile phase is needed for the electrolytes when using an elec-... 

MK Balz, E Schulte, H-P Thier. Simultaneous determination of a-tocopheryl acetate, tocopherols and tocotrienols by HPLC with fluorescence detection in foods. Fat Sci Techno] 95 215-220, 1993. 

The distribution and elimination of SARA from tissue of juvenille channel catfish was evaluated as the loss of radioactivity of SARA Cl 4 in liver, kidney, skin, and skinless fillet. An HPLC with fluorescence detection, in-line radiometric detection, and gradient elution was applied (203). The pharmacokinetics of ENRO in the milk of lactating cows was studied by HPLC, and it was found that a marked proportion of ENRO was metabolized to CIPRO (204). 

Since BAs occurring in food do not exhibit satisfactory absorbance or fluorescence in the visible or ultraviolet range, chemical derivatization, either pre- (35-37) or postcolumn (38), is usually used for their detection in HPLC. The most frequently employed reagents for precolumn derivatization are fluorescamine, aminoquinolyl-lV-hydroxysuccinimidyl carbamate (AQC) (39, 40), 9-fluorenylmethyl chloroformate (FMOC) (41-43), 4-dimethylaminoazobenzene-4 -sul-fonyl chloride (dabsylchloride, DBS) (44), N-acetylcysteine (NAC) (45,46), and 5-dimethyl-amino-1-naphthalene-1-sulfonyl chloride (dansylchloride, DNS) (47,48), phthalaldehyde (PA), and orf/to-phthaldialdehyde (OPA) (49-51), together with thiols such as 3-mercaptopropionic acid (MPA) (37) and 2-mercaptoethanol (ME) (35,49). 

A qualitative thin-layer chromatography method has been described by Kleef et al. [27] for the detection of rocuronium bromide and its metabolites in biological samples. This method was developed to confirm the identity of rocuronium bromide and its metabolites prior to their determination using HPLC coupled with fluorescence detection. In this method, the dried residue from the extraction process was dissolved in 0.05 ml of 0.01 M HC1. The stationary phase used was silicagel plates, that were developed in a mobile phase consisting of 2% solution of Nal in 2-propanol. The elution process was run for 4 h, and after the elution... 

Increased sensitivity of detection in HPLC can often be gained by monitoring the column effluent for intrinsic protein fluorescence by excitation... 

The main attraction of fluorescence detection for HPLC is that for strongly fluorescent molecules, it can offer limits of detection two or three orders of magnitude lower than UV absorbance methods. The reason for this lies in the difference in the nature of the measurements. In a UV absorbance detector the photodetector is constantly illuminated at a high... 

As regards the sensitivity of the MS detection in HPLC analysis of flavonoids, this technique has proved to be the most sensitive as compared to UV and fluorescence detection. A very comprehensive comparison of the four detection systems—UV, fluorescence, and two MS systems [atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI)]—for the determination of the previously identified 3, 4, 5 -trimethoxyflavone is presented in Table 1. Fluorescence detection is 10 times more sensitive than UV detection, whereas MS detection is 50 times more sensitive than UV detection and 5 times more sensitive than fluorescence detection. 

International Organization of Vine and Wine (2006). Measuring ochratoxin A in wine after going through an immunoaffinity column and HPLC with fluorescence detection, In Compendium of international methods of wine and must analysis. International Organization of Vine and Wine, Vol. 2. 

A wide variety of other esters has been made, or may potentially be accessible, by the type of procedure outlined in this section. To give a couple of examples, the naphthacyl esters were made for improved detectability in HPLC analysis in the same way as the phenacyl esters mentioned earlier [102], the fluorescent esters of 4-methyl-7-methoxycoumarin were made from the 4-bromomethyl-7-methoxycoumarin by the potassium salt procedure [103] and anthrylmethyl esters were obtained by an extractive alkylation procedure [104]. 

Z. Loukou, A. Zotou, A comparative survey of the simultaneous ultraviolet and fluorescence detection in the RP-HPLC determination of dansylated biogenic amines in alcoholic beverages, Chromatographia 2003, 58, 579. 

O Flaherty, B. Fluorescence detection. In A Practical Guide to HPLC Detection Parriott, D., Ed. Academic Press San Diego, CA, 1993 111 139. 

Because gabapentin lacks chromophores, UV detection is difficult. Therefore, analysis was usually carried out by precolumn derivatization followed by RPLC with UV or fluorescence detection. In contrast, the method of Jia et al. [156] does not require any derivatization. In their study, the performance of CAD and ELSD were compared on four different HILIC columns. It was found that the sensitivity achieved by ELSD was comparable to that with CAD and both superior to UV detection in HlLlC mode. However, employing a conventional CIS column, CAD was about 25-times more sensitive than ELSD. Jia et al. also introduced HPLC-CAD analysis of gabapentin in rat serum and mine, which is the first report of a successful application of CAD in such matrices [157],... 

In recent years, RP-HPLC with UV (or fluorescence) detection methods have also been successfully used for the determination of B vitamins in honey. Vinas et al. (2004a, 2004b) proposed quantifying vitamins B2 and Bg with chromatographic methods using fluorescence detection. In their protocol for B2, the stationary phase is an end-capped amide-based column and the mobile phase is a 10 90 v/v acetonitrile/phosphate buffer (pH = 5). The protocol for Bg is... 

---