Highly sensitive spectrofluorimetric method for rapid determination of orciprenaline in biological fluids and pharmaceuticals.

Orciprenaline sulphate (ORP) is a direct-acting sympathomimetic with mainly beta-adrenoceptor stimulant activity. It is used as a bronchodilator in the management of reversible airway obstruction. For the first time, a rapid highly sensitive spectrofluorimetric method is described that is relied on measuring the fluorescence spectra of ORP at acidic pH and without addition of any chemical reagents. The relative fluorescence intensity was measured at 310 nm and after excitation at 224 nm. ORP native fluorescence was calibrated in both water and acetonitrile as diluting solvents. The method was designed to estimate the drug in miscellaneous matrices with high accuracy and precision. Linear ranges of calibration curves were 30.0-400.0 ng/ml and 10.0-240.0 ng/ml in water and acetonitrile, respectively. The detection limits were calculated and reached as low as 3.3 and 3.1 ng/ml, respectively, representing the ultra-sensitivity of the proposed method. This result permitted application of this method for spiked human plasma and urine and was used as a preliminary investigation with good percentage recovery (89.4-106.8%). The application was further extended to analyse ORP in its pharmaceutical formulations. The method was validated in compliance with International Council of Harmonization (ICH) Guidelines.

Purification of pharmaceutical preparations using thin-layer chromatography to obtain mass spectra with Direct Analysis in Real Time and accurate mass spectrometry.

Forensic analysis of pharmaceutical preparations requires a comparative analysis with a standard of the suspected drug in order to identify the active ingredient. Purchasing analytical standards can be expensive or unattainable from the drug manufacturers. Direct Analysis in Real Time (DART™) is a novel, ambient ionization technique, typically coupled with a JEOL AccuTOF™ (accurate mass) mass spectrometer. While a fast and easy technique to perform, a drawback of using DART™ is the lack of component separation of mixtures prior to ionization. Various in-house pharmaceutical preparations were purified using thin-layer chromatography (TLC) and mass spectra were subsequently obtained using the AccuTOF™- DART™ technique. Utilizing TLC prior to sample introduction provides a simple, low-cost solution to acquiring mass spectra of the purified preparation. Each spectrum was compared against an in-house molecular formula list to confirm the accurate mass elemental compositions. Spectra of purified ingredients of known pharmaceuticals were added to an in-house library for use as comparators for casework samples. Resolving isomers from one another can be accomplished using collision-induced dissociation after ionization. Challenges arose when the pharmaceutical preparation required an optimized TLC solvent to achieve proper separation and purity of the standard. Purified spectra were obtained for 91 preparations and included in an in-house drug standard library. Primary standards would only need to be purchased when pharmaceutical preparations not previously encountered are submitted for comparative analysis. TLC prior to DART™ analysis demonstrates a time efficient and cost saving technique for the forensic drug analysis community. Copyright © 2011 John Wiley & Sons, Ltd.

Multiresidue analysis of beta-agonists in bovine and porcine urine, feed and hair using liquid chromatography electrospray ionisation tandem mass spectrometry.

The use of beta-agonists as growth promoters in cattle breeding is forbidden in many countries for reasons of fair trade and consumer protection. In recent years the use of liquid chromatography (LC) tandem mass spectrometry (MS/MS) has been shown to be the method of choice for the control of beta-agonists. In this study an LC-MS/MS multiresidue analysis method is presented for trace analysis of 22 beta-agonists. A truly generic concept has been designed based on mixed-mode solid-phase extraction and positive electrospray ionisation LC-MS/MS operated in the multiple reaction monitoring mode. This method allows application to a wide variety of sample matrices such as urine, feed and hair, following minor modifications to the analysis procedure only. The method features fit-for-purpose sensitivity in urine as shown by CCalpha and CCbeta values of less than 0.2 and less than 0.5 microg/l respectively, for all beta-agonists studied (terbutaline and reproterol, less than 0.3 and less than 1.0 respectively). Similar but semiquantitative application to feed and hair showed CCbeta values of less than 10.0 and less than 5.0 microg/kg, respectively. A further simplification and improvement is demonstrated using Ultra Performance LC (UPLC) and fast-switching MS/MS. The successful validation of this method following the latest EU requirements and its application to real samples demonstrate that a new versatile tool has been achieved for veterinary control of beta-agonists.

A novel chemiluminescence method for the determination of orciprenaline based on ferricyanide-rhodamine 6G.

A novel flow injection chemiluminescence method for the determination of orciprenaline was developed. The method is based on the chemiluminescence (CL) reaction of orciprenaline with potassium ferricyanide in sodium hydroxide medium, sensitized by the fluorescent dye rhodamine 6G. The proposed procedure allows quantitation of orciprenaline in the concentration range 0.01-1.2 microg/mL, with a detection limit of 7.2 x 10(-3) microg/mL. The relative standard deviation (RSD) is 2.7% for 0.1 microg/mL orciprenaline (n = 9). The sampling frequency was calculated at approximately 120/h. The method was successfully applied to the determination of orciprenaline in pharmaceutical preparations. A brief discussion on the possible CL reaction mechanism is presented.

Conductimetric determination of reproterol HCl and pipazethate HCl and salbutamol sulphate in their pharmaceutical formulations.

A simple and sensitive conductimetric method for the determination of salbutamol sulphate and reproterol and pipazethate hydrochlorides is presented based on their ion associates with phosphotungstic and phosphomolybdic acids. The effect of solvent, molar ratio, reagent concentration and temperature were studied, and the solubility products of the formed ion associates were calculated. The method was applied to the determination of the drugs in their pure state or pharmaceutical preparations with mean recovery values of 99.82-100.54, 99.75-100.12 and 99.95-100.40%, and coefficient of variation 0.28-0.52, 0.16-0.36 and 0.19-0.33 for salbutamol sulphate, reproterol HCl and pipazathate HCl, respectively.

Screening of beta-2 agonists and confirmation of fenoterol, orciprenaline, reproterol and terbutaline with gas chromatography-mass spectrometry as tetrahydroisoquinoline derivatives.

A new method for a comprehensive screening and confirmation of beta-2 agonists in human urine is presented based on gas chromatography-low-resolution mass spectrometry (GC-MS) using electron impact ionisation (EI). After hydrolysis of the conjugates with beta-glucuronidase/arylsulfatase a derivatisation step with formaldehyde converts fenoterol, orciprenaline, reproterol and terbutaline to one derivative, a tetrahydroisoquinoline, while the other beta-2 agonists remain unchanged. Liquid-liquid extraction and trimethylsilylation follow. The tetrahydroisoquinoline derivatives show good gas chromatographic and mass spectrometric behaviour. The detection limit of these four beta-2 agonists in the screening using low-resolution mass spectrometry is 10 ng/ml of urine. The other beta-2 agonists are detected as parent compounds with the same recovery after sample preparation with and without formaldehyde. The EI mass spectra of the tetrahydroisoquinoline derivatives are presented.

Coulometric determination of some antiasthmatics.

A simple and rapid method for the assay of microquantities of fenoterol hydrobromide, orciprenaline sulphate and terbutaline sulphate in pure state and various pharmaceutical formulations, is presented. The method is based on the coulometric titration of the investigated compounds with electrogenerated chlorine in the presence of methyl orange as indicator. The method requires a simple apparatus and gives accurate and reproducible results.

Chemical stabilities of isoetharine hydrochloride, metaproterenol sulphate and terbutaline sulphate after mixing with normal saline for respiratory therapy.

The chemical stabilities of isoetharine hydrochloride inhalation solution, metaproterenol sulphate inhalation solution and terbutaline sulphate injection, after diluting 1 in 10 with sodium chloride 0.9% injection were studied. On storing the solutions in amber-coloured syringes, they were stable for at least 120 days at 5 degrees C. At 25 degrees C they were also stable for 120 days except that isoetharine solution discoloured and lost 7.8% of its potency after 90 days of storage. There was a new peak in the chromatogram from the decomposition product. All other solutions remained clear for 120 days at both temperatures. The initial and final pH values were similar except that after 120 days at both temperatures. The initial and final pH values were similar except that after 120 days at 25 degrees C, the pH value of terbutaline solution had increased from 4.9 to 5.4.

Colorimetric determination of terbutaline sulfate and orciprenaline sulfate via nitrosation and difference spectrophotometry.

A colorimetric method is proposed for determination of terbutaline sulfate, orciprenaline sulfate, and their dosage forms. The suggested method depends on nitrosation of the 2 drugs by using sodium nitrite and hydrochloric acid. Addition of sodium hydroxide increases the intensity of the color developed. The difference between absorption values measured in acid and alkaline media is taken as a measure of concentration. Variables were carefully studied and optimized. Results for both compounds adhered to Beer's law over the range 2-28 micrograms/mL. The method has proved to be accurate and precise for analysis of pharmaceutical dosage forms.

Determination of dissociation constants of 5-[1-hydroxy-2-[(1-methylethyl)amino]ethyl]-1-3-benzenediol.

The microdissociation constants of 5-[1-hydroxy-2-[ (methylethyl)amino]ethyl]-1-3-benzenediol have been determined by means of spectrophotometric and spectrofluorometric techniques. Experimental data were analyzed using the linear regression method. Values of 8.90, 10.0, 10.25, 9.15, and 12.5 have been found for the microdissociation constants.

Detection of sulfur dioxide in bronchodilator aerosols.

Several bronchodilator drugs commonly used in respiratory therapy contain sodium metabisulfite as an antioxidant preservative. When aerosolized, these agents may release the irritant gas SO2 as a result of bisulfite decomposition. We found that agents that contain bisulfites generated SO2 concentrations of 2.0 ppm and greater, while bronchodilator solutions without bisulfite did not. Such levels are known to induce or to exacerbate asthmatic symptoms. Levels of SO2 were higher when solutions were nebulized with compressed air from a tank or electric compressor than when they were nebulized from a hand-bulb nebulizer. No significant lot-to-lot variations were found in the solutions tested.

Simple colorimetric method for determination of metaproterenol sulfate in pharmaceuticals.

A simple colorimetric method is described for the determination of metaproterenol sulfate (orciprenaline sulfate). The method is based on measurement of a colored species formed when metaproterenol sulfate is treated with diazotized dapsone, p-nitroaniline, or benzocaine at room temperature, followed by treatment with an aqueous solution of trimethylamine in the case of benzocaine. Compounds such as starch, talc, and common excipients do not interfere in the reaction. Statistical validation showed that the method was precise and accurate. The results agree well with those obtained by other methods reported in the literature.

Biotransformation of reproterol in the intestinal tract of the rat.

The major metabolite of reproterol (Broncho-spasmin) in rat feces is 2-[3-theophyllinyl(7)-propyl]-4,6,8-trihydroxy-1,2,3,4-tetrahydroisoquinoline. It was also found in the bile of orally or intravenously dosed rats in the form of glucuronides. The biotransformation of an oral dose of reproterol to this metabolite occurred mostly in the cecum-colon section of the intestinal tract. Experiments in antibiotic treated rats showed no significant effect of the treatment on the extent of metabolite formation. This metabolite was also formed by incubation of reproterol with cecum-colon homogenates under aerobic as well as anaerobic conditions. The pharmacological action after an oral dose must be attributed to reproterol absorbed as intact form, since the metabolite is inactive.