Selective determination of amino acids using flow injection analysis coupled with chemiluminescence detection

The determination of the amino acids proline, histidine, tyrosine, arginine, phenylalanine and tryptophan using flow injection analysis (FIA) with chemiluminescence detection is described. Proline was the only amino acid to exhibit chemiluminescence with the tris(2,2-bipyridyl)ruthenium(III) reaction at pH 10. While, histidine was found to selectively enhance the reaction of luminol with Mn(II) salts in a basic medium. Acidic potassium permanganate chemiluminescence was able to selectively determine tyrosine at pH 6.75. Low pressure separations using a C18 guard column allowed the simultaneous determination of tyrosine and tryptophan or phenylalanine and tryptophan with acidic potassium permanganate and copper(II)–amino acid–hydrogen peroxide chemiluminescence, respectively. Precision for each method was less than 3.9% (R.S.D.) for five replicates of a standard (1×10−5 M) and the detection limits ranged between 4×10−9 and 7×10−6 M. Preliminary investigations revealed that the methodology developed was able to selectively determine the individual amino acids in an equimolar mixture of the 20 naturally occurring amino acids.

Determination of glyphosate mono-isopropylamine salt in process samples using flow injection analysis with tris(2,2`-bipyridyl)ruthenium(II) chemiluminescence detection

The mono-isopropylamine salt of glyphosate was selectively determined directly in industrial and commercial formulations using flow injection analysis with tris(2,2′-bipyridyl)ruthenium(II) chemiluminescence detection without the need for separation. Glyphosate and its mono-isopropylamine salt furnished detection limits of 7×10⁻⁹ and 3.5×10⁻¹⁰ M and relative standard deviations of 0.4% at 1×10⁻⁷ M and 0.8% at 5×10⁻⁸ M, respectively. The methodology is robust and reliable with samples subjected only to aqueous dilution prior to analysis.

Determination of proline in wine using flow injection analysis with tris(2,2`-bipyridyl)ruthenium(II) chemiluminescence detection

Flow injection methodology is described for the determination of proline in red and white wines using tris(2,2′-bipyridyl)ruthenium(II) chemiluminescence detection. Selective conditions were achieved for proline at pH 10, while other amino acids and wine components did not interfere. The precision of the method was less than 1.00% (R.S.D.) for five replicates of a standard (4 × 10⁻⁶ M) and the detection limit was 1 × 10⁻⁸ M. The level of proline in white and sparkling wines using the developed methodology was equivalent to those achieved using HPLC-FMOC amino acid analysis. SPE removal of phenolic material was required for red wines to minimize Ru(bipy)₃³⁺ consumption and its associated effect on accuracy.

Determination of psilocin and psilocybin using flow injection analysis with acidic potassium permanganate and tris(2,2’-bipyridyl) ruthenium(II) chemiluminescence detection respectively

A simple, rapid and sensitive method for the determination of psilocin and psilocybin is described. This is the first report on the determination of psilocin and psilocybin using flow injection analysis with acidic potassium permanganate and tris(2,2′-bipyridyl)ruthenium(II) chemiluminescence. The limits of detection (signal-to-noise ratio = 3) are 9 × 10⁻¹⁰ M and 3 × 10⁻¹⁰ M for psilocin and psilocybin, respectively.A concise synthetic route for psilocin in three steps from readily available starting materials is also described. The structures were elucidated on the basis of spectroscopic data.

The determination of morphine in the larvae of calliphora stygia using flow injection analysis and HPLC with chemiluminescence detection

Selective determination of morphine in the larvae of Calliphora stygia (Fabricius) (Diptera: Calliphoridae) using acidic potassium permanganate chemiluminescence detection coupled with flow injection analysis and high-performance liquid chromatography (HPLC) is described. Larvae of C. stygia were reared on minced meat substrates that had been spiked with varying concentrations of morphine. Morphine concentrations were chosen to reflect typical levels in human tissues from opiate overdose victims. After maturing on substrates, larvae were analyzed for the presence of morphine using chemiluminescence detection coupled to flow injection analysis and a rapid HPLC method. Analysis of the larval matrix by flow injection analysis with chemiluminescence detection indicated the presence of interferants capable of generating chemiluminescence. A rapid chromatographic separation with a monolithic column allowed selective determination of morphine in larvae using postcolumn chemiluminescence detection. Larvae of C. stygia reared on substrates containing morphine at concentrations of 500 and 1000 ng/g did not sequester morphine at detectable concentrations. Larvae reared on substrates containing morphine concentrations of 2500, 5000, and 10,000 ng/g tested positive for the drug at concentrations of 765, 2720, and 3010 ng/g, respectively.

A sensitive procedure for the rapid determination of arsenic(III) by flow injection analysis and chemiluminescence detection

A novel chemiluminescence flow injection procedure for the determination of As(III) in aqueous samples is described. The method involves injection of As(III) samples into a 1% (m/v) sodium hexametaphosphate in 0.02 M H₂SO₄ carrier stream, which then merges at a Y-piece with a reagent stream consisting of potassium permanganate (5.0 × 10⁻⁵ M) made up in the acidic sodium hexametaphosphate carrier solution. The chemiluminescence intensity of the resulting reaction mixture was measured at a photomultiplier tube operated at a voltage of 0.93 kV. Under optimized conditions, the method is characterised by a linear range from 0.5 to 5.0 μg l⁻¹, a detection limit of 0.3 μg l⁻¹ and a sampling frequency of 150 h⁻¹. The effects of common anionic and cationic interferences were investigated, and it was found that the only ions to cause serious interference were those which react with potassium permanganate, namely sulphide, iodide and ferrous.

Chemiluminescence detection flow cells for flow injection analysis and high-performance liquid chromatography

We have examined a range of new and previously described flow cells for chemiluminescence detection. The reactions of acidic potassium permanganate with morphine and amoxicillin were used as model systems representing the many fast chemiluminescence reactions between oxidising agents and organic analytes, and the preliminary partial reduction of the reagent was exploited to further increase the rates of reaction. The comparison was then extended to high-performance liquid chromatography separations of α- and β-adrenergic agonists, with permanganate chemiluminescence detection. Flow cells constructed by machining novel channel designs into white polymer materials (sealed with transparent films or plates) have enabled improvements in mixing efficiency and overall transmission of light to the photodetector.

Comparison of soluble manganese(IV) and acidic potassium permanganate chemiluminescence detection using flow injection and sequential injection analysis for the determination of ascorbic acid in vitamin C tablets

The limits of detection (3s) for ascorbic acid were 5×10⁻⁸ M with acidic potassium permanganate using both flow injection analysis (FIA) and sequential injection analysis (SIA) whereas the soluble manganese(IV) afforded 1×10⁻⁸ M and 5×10⁻⁹ M for FIA and SIA, respectively. Determinations of ascorbic acid in Vitamin C tablets were achieved with minimal sample pretreatment using a standard additions calibration and gave good agreement with those of iodimetric titration.

Monitoring the total phenolic/antioxidant levels in wine using flow injection anaylsis with acidic potassium permanganate chemiluminescence detection

Flow injection methodology is described for the estimation of the total phenolic content of wine using acidic potassium permanganate chemiluminescence detection. Selected simple phenolic compounds including quercetin, rutin, catechin, epicatechin, ferulic acid, caffeic acid, gallic acid, 4-hydroxycinnamic acid and vanillin elicited analytically useful chemiluminescence with detection limits ranging between 4×10⁻¹⁰ and 7×10⁻⁷ M. A comparison between the chemiluminescence methodology and other total phenol/antioxidant assays, used by the food and beverage industry, resulted in a good correlation. The chemiluminescence detection was found to be selective with minimal interferences being observed from the non-phenolic components in wine. Analysis of 12 different wines showed that the chemiluminescence method was a rapid way to estimate their antioxidant or total phenolic content.

Detection of pyrrolizidine alkaloids using flow analysis with both acidic potassium permanganate and tris(2,2’-bipyridyl)ruthenium(II) chemiluminescence

For the first time, analytically useful chemiluminescence was elicited from the reactions of the pyrrolizidine alkaloids. Heliotrine, retronecine, supinine, monocrotaline and echinatine N-oxide yielded chemiluminescence upon reaction with tris(2,2′-bipyridyl)ruthenium(II) whilst lasiocarpine, its N-oxide and supinine elicited light upon reaction with acidic potassium permanganate. Detection limits for heliotrine were 1.25 × 10⁻⁷ M and 9 × 10⁻⁹ M for tris(2,2′-bipyridyl)ruthenium(III) perchlorate with flow injection analysis (FIA) and the silica-immobilised reagent (4-[4-(dichloromethylsilanyl)-butyl]-4′-methyl-2,2′-bipyridine)bis(2,2′-bipyridyl)ruthenium(II) with sequential injection analysis (SIA), respectively. Lasiocarpine was detectable at 1.4 × 10⁻⁷ M using acidic potassium permanganate with FIA. Additionally, the silica-immobilised reagent was optimised with respect to the oxidant (ammonium ceric nitrate) concentration and the aspiration times which afforded a detection limit for codeine of 5 × 10⁻¹⁰ M using SIA.

Design of LabVIEW® - based software for the control of sequential injection analysis instrumentation for the determination of morphine.

LabVIEW^®-based software for the automation of a sequential injection analysis instrument for the determination of morphine is presented. Detection was based on its chemiluminescence reaction with acidic potassium permanganate in the presence of sodium polyphosphate. The calibration function approximated linearity (range 5 × 10 ^-10 to 5 × 10 ^-6M) with a line of best fit of y = 1.05 x + 8.9164 (R² = 0.9959), where y is the log₁₀ signal (mV) and x is the log₁₀ morphine concentration (M). Precision, as measured by relative standard deviation, was 0.7% for five replicate analyses of morphine standard (5 × 10^-8_M). The limit of detection (3 σ) was determined as 5 × 10^-11 M morphine.

A screening test for heroin based on sequential injection analysis with dual-reagent chemiluminescence detection

A sequential injection analysis procedure with dual-reagent chemiluminescence detection was applied to the screening of street drug seizure samples for the presence of heroin. The chemiluminescence reagents (acidic potassium permanganate and tris(2,2′-bipyridine)ruthenium(III)) were aspirated from either side of a sample aliquot that was sufficiently large to prevent interdispersion of the reagent zones, and therefore two different chemical reactions could be performed simultaneously at either end of the sample zone. The presence of heroin in seizure samples was indicated by a strong response with the tris(2,2′-bipyridine)ruthenium(III) reagent and confirmed by a significant increase in the response with the permanganate reagent when the sample was treated with sodium hydroxide to hydrolyse the heroin to morphine. Nicomorphine (a morphine-derived pharmaceutical) was synthesised and tested under the same conditions. The responses with the permanganate reagent were similar to those for heroin, which supports the proposed chemical basis for the test. However, the responses with tris(2,2′-bipyridine)ruthenium(III) were far lower for nicomorphine than heroin (approximately 5-fold for the samples that had not been hydrolysed).

Precision milled flow-cells for chemiluminescence detection

Novel flow-cells with integrated confluence points and reaction channels designed for efficient mixing of fast chemiluminescence systems were constructed by machining opposing sides of a polymer chip and sealing the channels with transparent epoxy-acetate films. A hole drilled through the chip provided the conduit from the confluence point on one side to the centre of the reaction zone on the other side, allowing rapid presentation of the reacting mixture to the photodetector. The effectiveness of each flow-cell was evaluated by comparing the chemiluminescence intensity using flow injection analysis methodology, and examining the distribution of light emanating from the reaction zone (captured by photography in a dark room) when the reactants were continuously merged. Although previously reported chemiluminescence detectors constructed by machining channels into polymers have almost exclusively been prepared using transparent materials, we obtained far greater emission intensities using an opaque white chip with a thin transparent seal, which minimised the loss of light through surfaces not exposed to the photomultiplier tube. Furthermore, this approach enabled the exploration of reactor designs that could not be incorporated in traditional coiled-tubing flow-cells.

3D-printed and CNC milled flow-cells for chemiluminescence detection

Herein we explore modern fabrication techniques for the development of chemiluminescence detection flow-cells with features not attainable using the traditional coiled tubing approach. This includes the first 3D-printed chemiluminescence flow-cells, and a milled flow-cell designed to split the analyte stream into two separate detection zones within the same polymer chip. The flow-cells are compared to conventional detection systems using flow injection analysis (FIA) and high performance liquid chromatography (HPLC), with the fast chemiluminescence reactions of an acidic potassium permanganate reagent with morphine and a series of adrenergic phenolic amines.