Spectrometer Operating Research Articles

Laboratory and astronomical discovery of the cyanovinyl radical H2CCCN

We report the first laboratory and interstellar detection of the α-cyano vinyl radical (H2CCCN). This species was produced in the laboratory by an electric discharge of a gas mixture of vinyl cyanide, CH2CHCN, and Ne. Its rotational spectrum was characterized using a Balle-Flygare narrowband-type Fourier-transform microwave spectrometer operating in the frequency region of 8–40 GHz. The observed spectrum shows a complex structure due to tunneling splittings between two torsional sublevels of the ground vibronic state, 0+ and 0−, derived from a large-amplitude inversion motion. In addition, the presence of two equivalent hydrogen nuclei makes it necessary to discern between ortho- and para-H2CCCN. A least-squares analysis reproduces the observed transition frequencies with a standard deviation of ca. 3 kHz. Using the laboratory predictions, this radical was detected in the cold dark cloud TMC-1 using the Yebes 40 m telescope and the QUIJOTE1 line survey. The 40, 4-30, 3 and 50, 5-40, 4 rotational transitions, composed of several hyperfine components, were observed in the 31.0–50.4 GHz range. Adopting a rotational temperature of 6 K, we derived a column density of (1.4±0.2)×1011 cm−2 and (1.1±0.2)×1011 cm−2 for ortho-H2CCCN and para-H2CCCN, respectively. The reaction of C + CH3CN emerges as the most likely route to H2CCCN in TMC-1, and possibly that of N + CH2CCH as well.

Low Barrier Methyl Internal Rotations and 14N Quadrupole Coupling in the Microwave Spectrum of 2,4-Dimethylthiazole.

The microwave spectrum of 2,4-dimethylthiazole was recorded using a pulsed molecular jet Fourier-transform microwave spectrometer operating in the frequency range from 2.0 to 26.5 GHz. Torsional splittings into quintets were observed for all rotational transitions due to internal rotations of two inequivalent methyl groups. Hyperfine structures arising from the nuclear quadrupole coupling of the 14N nucleus were fully resolved. The microwave spectra were analyzed using the modified version of the XIAM code and the BELGI-Cs-2Tops-hyperfine code. The barriers to methyl internal rotation of the 4- and 2-methyl groups were determined to be 396.707(25) cm-1 and 19.070(58) cm-1, respectively. The very low barrier hindering the 2-methyl torsion was a challenge for the spectral analysis and modeling, and separately fitting the five torsional species together with combination difference loops was the key for a successful assignment. The methyl torsional barriers were compared with those of other thiazole derivatives, showing the influence of the methyl group position on the barrier height. The experimental results were supported by quantum chemical calculations.

Development of Methods for the Synthesis of Malathion-D<sub>6</sub>, Chlorophos-D<sub>6</sub>, and Dichlorophos-D<sub>6</sub> for Use as Internal Standards in the Analysis of Medicinal Plant Raw Materials and Herbal Medicinal Products

Scientific relevance. Deuterated analogues of test substances, when used as internal standards, enable analysts to determine traces of residual pesticides in a medicinal plant raw material by gas or liquid chromatography coupled with mass spectrometry. This prompts the search for an undemanding method for the synthesis of deuterated reference standards, with the ultimate goal of expanding the range of reference standards produced in Russia.Aim. The study aimed to synthesise deuterated reference standards (malathion-D6, chlorophos-D6, dichlorophos-D6, and their precursors) and determine their spectral characteristics using 1Н, 2Н, 13С and 31Р nuclear magnetic resonance (NMR) spectroscopy and high-resolution mass spectrometry.Materials and methods. The study involved methanol-D4, phosphorus pentasulfide, phosphorus trichloride, maleic acid diethyl ether, triethylamine, hydroquinone, and chloral hydrate. The authors recorded 1H, 2H, 13C, and 31P NMR spectra using a QONE AS400 NMR spectrometer operating at a temperature of 298 °K and respective frequencies of 399.83, 61.38, 100.55, and 161.85 MHz. Mass spectra were obtained using an analytical system comprising an Agilent Infinity 1260 liquid chromatograph equipped with an automatic sample injector and coupled with a Thermo Scientific Orbitrap Fusion Lumos high-resolution mass-selective tandem spectrometer operating in electrospray ionisation mode.Results. The authors used existing methods for the synthesis of organophosphate pesticides as a basis and developed synthesis routes to produce the deuterated analogues malathion-D6, chlorophos-D6, and dichlorophos-D6. The synthesised compounds were identified by NMR spectroscopy and high-resolution mass spectrometry.Conclusions. The synthesised compounds are highly pure and suitable for use as internal standards in the determination of organophosphate pesticides in medicinal plant raw materials. The synthesis methods developed by the authors can be used to produce reference standards in Russia.

Rotational spectra and astronomical search of five ethynyl derivatives of propene

ABSTRACT The interstellar discovery of the five cyano derivatives of propene brings interest in analogous molecules that could be detected in similar astronomical environments. In this context, ethynyl-propene derivatives are good candidates to be observed in the Taurus Molecular Cloud (TMC-1), where other cyano and ethynyl derivatives of hydrocarbons such as ethane, allene, propene, and cyclopentadiene, among others, have been detected. We have investigated the pure rotational spectra of the five ethynyl-propene derivatives to enable their interstellar detections. These molecular species have been produced by an electric discharge and their rotational spectra were characterized using a Balle–Flygare narrow band-type Fourier-transform microwave spectrometer operating in the frequency region of 4-40 GHz. The spectral analysis allowed us to derive accurate spectroscopic parameters to obtain reliable frequency predictions for astronomical searches. We do not detect any of these species in TMC-1 but we derive upper limits for the abundances for the five isomers.

Rapid target screening and quantitative analysis of various environmental pollutants.

The presented analytical method demonstrates a straightforward approach for environmental applications based on laser diode thermal desorption (LDTD). The study aims to examine abilities to achieve environmentally relevant outcomes for different types of pollutants with a fast method following the green chemistry principle. Treatment of environmentally relevant sample matrix (river water) was limited to filtration with a cellulose filter. Samples fortified with analytes were spotted in a LazWell plate and dried before analysis. Samples thermally desorbed using LDTD were detected with Q Exactive hybrid high-resolution mass spectrometer operation in full-scan data-dependent acquisition mode (LDTD-FullMS-dd-MS/MS). LDTD-FullMS-dd-MS/MS exhibits the lowest quantification limits for anatoxin-A, atrazine, caffeine, methamphetamine, methylbenzotriazole, paracetamol, perfluorobutanoic acid, perfluorohexanoic acid, and perfluorooctanoic acid of between 0.10 and 1.0ng mL-1 in the environmentally relevant sample matrix. The developed method was successfully evaluated for different environmental pollutants and radically reduced sample treatment and time requirements for analysis and sample preparation.

Development of a liquid chromatography-tandem mass spectrometry (LC–MS/MS) method for characterizing pomegranate extract pharmacokinetics in humans

Pomegranate extracts standardized to punicalagins are a rich source of ellagitannins including ellagic acid (EA). Recent evidence suggests that gut microbiota-derived urolithin (Uro) metabolites of ellagitannins are pharmacologically active. Studies have evaluated the pharmacokinetics of EA, however, little is known about the disposition of urolithin metabolites (urolithin A (UA) and B (UB)). To address this gap, we developed and applied a novel ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) assay for the characterization of EA and Uro oral pharmacokinetics in humans. Subjects (10/cohort) received a single oral dose (250 or 1000 mg) of pomegranate extract (Pomella® extract) standardized to contain not less than 30 % punicalagins, < 5 % EA, and not less than 50 % polyphenols. Plasma samples, collected over 48 h, were treated with β-glucuronidase and sulfatase to permit comparison between unconjugated and conjugated forms of EA, UA and UB. EA and urolithins were separated by gradient elution (acetonitrile/water, 0.1 % formic acid) using a C18 column connected to a triple quadrupole mass spectrometer operating in the negative mode. Conjugated EA exposure was ∼5–8-fold higher than unconjugated EA for both dose groups. Conjugated UA was readily detectable beginning ∼8 h post-dosing, however, unconjugated UA was detectable in only a few subjects. Neither form of UB was detected. Together these data indicate EA is rapidly absorbed and conjugated following oral administration of Pomella® extract. Moreover, UA’s delayed appearance in the blood, primarily in the conjugated form, is consistent with gut microbiota-mediated metabolism of EA to UA, which is then rapidly converted to its conjugated form.

Two-photon absorption spectrometers for near infrared

We present a Silicon-based Charge-Coupled Device (Si-CCD) sensor applied as a cost-effective spectrometer for femtosecond pulse characterization in the Near Infrared region in two different configurations: two-Fourier and Czerny-Turner setups. To test the spectrometer's performance, a femtosecond Optical Parametric Oscillator with a tuning range between 1100 and 1700nm and a femtosecond Erbium-Doped Fiber Amplifier at 1582nm were employed. The nonlinear spectrometer operation is based on the Two-Photon Absorption effect generated in the Si-CCD sensor. The achieved spectrometer resolution was 0.6 ± 0.1nm with a threshold peak intensity of 2×106Wcm2. An analysis of the nonlinear response as a function of the wavelength, the response saturation, and the criteria to prevent it are also presented.

Simultaneous determination of UDCA and its major metabolites in human plasma with surrogate matrix by a rapid and specific LC-MS/MS method

A rapid, convenient, and specific liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous quantification of ursodeoxycholic acid (UDCA), and its major metabolites, glycoursodeoxycholic acid (GUDCA) and tauroursodeoxycholic acid (TUDCA) in human plasma. Methanol was chosen as surrogate matrix for preparation the calibrators to establish calibration curves. Isotope internal standard was used for each analyte. After plasma samples were deproteinized with methanol, the post-treatment samples were analyzed on a ZORBAX SB-C18 column (2.1 × 50 mm, 1.8 μm) with 2 mM ammonium acetate and acetonitrile as mobile phase at a flow rate of 0.5 mL/min. Detection was performed on a triple quadrupole mass spectrometer operating in multiple reaction monitoring (MRM) employing negative ESI interface using API5500 triple quadrupole tandem mass spectrometer system, with the transitions set at m/z 391.4 → m/z 391.4, m/z 448.3 → m/z 73.9, m/z 498.4 → m/z 80.1, m/z 395.3 → m/z 395.3, m/z 453.3 → m/z 74.0, and m/z 503.2 → m/z 79.9 for UDCA, GUDCA, TUDCA, UDCA-d4, GUDCA-d5, and TUDCA-d5, respectively. The calibration curve ranges were 5.00–2500 ng/mL for UDCA and GUDCA and 0.500–250 ng/mL for TUDCA. The intra- and inter-day precision was within 7.00% in terms of relative standard deviation (RSD%) and the accuracy within 11.75% in terms of relative error. The selectivity, sensitivity, extraction recovery, matrix effect, dilution reliability, and stability were within the acceptable range. The method was successfully applied to a pharmacokinetic study in 12 healthy Chinese volunteers after oral administration of 250 mg UDCA.

Versatile OSCAT time-domain THz spectrometer.

We report on a compact and versatile time-domain spectrometer operating in the THz spectral region from 0.2 to 2.5 THz based on ultrafast Yb:CALGO laser and photo-conductive antennas. The spectrometer operates with the optical sampling by cavity tuning (OSCAT) method based on laser repetition rate tuning, which allows at the same time the implementation of a delay-time modulation scheme. The whole characterization of the instrument is presented and compared to the classical THz time-domain spectroscopy implementation. THz spectroscopic measurements on a 520-μm thick GaAs wafer substrate together with water vapor absorption measurements are also reported to further validate the instrument capabilities.

Rate constants for saturation-recovery EPR and ELDOR of 14N-Spin labels

Saturation-recovery (SR)-EPR can determine electron spin–lattice relaxation rates in liquids over a wide range of effective viscosity, making it especially useful for biophysical and biomedical applications. Here, I develop exact solutions for the SR-EPR and SR-ELDOR rate constants of 14N-nitroxyl spin labels as a function of rotational correlation time and spectrometer operating frequency. Explicit mechanisms for electron spin–lattice relaxation are: rotational modulation of the N-hyperfine and electron-Zeeman anisotropies (specifically including cross terms), spin-rotation interaction, and residual frequency-independent vibrational contributions from Raman processes and local modes. Cross relaxation from mutual electron and nuclear spin flips, and direct nitrogen nuclear spin–lattice relaxation, also must be included. Both the latter are further contributions from rotational modulation of the electron-nuclear dipolar interaction (END). All the conventional liquid-state mechanisms are defined fully by the spin-Hamiltonian parameters; only the vibrational contributions contain fitting parameters. This analysis gives a firm basis for interpreting SR (and inversion recovery) results in terms of additional, less standard mechanisms.

Microbial degradation products of lurasidone and their significance in postmortem toxicology.

Recent research reported that lurasidone degrades in unpreserved ante-mortem human whole blood inoculated with microorganisms known to dominate postmortem blood specimens. In vitro degradation occurred at a similar rate to risperidone, known to degrade in authentic postmortem specimens until below analytical detection limits. To identify the lurasidone degradation products formed, an Agilent 6520 liquid chromatograph quadrupole-time-of-flight mass spectrometer (LC-QTOF-MS) operating in auto-MS/MS mode was used. Numerous degradation products not previously reported in prior in vitro or in vivo pharmacokinetic studies or forced degradation studies were detected. Accurate mass data, mass fragmentation data, acetylation experiments, and a proposed mechanism of degradation analogous to risperidone supports initial identification of the major degradation product as N-debenzisothiazole-lurasidone (calculated m/z [M + H]+ = 360.2646). A standard was unavailable to conclusively confirm this identification. Retrospective data analysis of postmortem cases involving lurasidone identified the presence of the major degradation product in four of six cases where lurasidone was also detected. This finding is significant for toxicology laboratories screening for this drug in postmortem casework. The major postmortem lurasidone degradation product has consequently been added to the LC-QTOF-MS drug screen at Forensic Science SA (FSSA) to indicate postmortem lurasidone degradation in authentic postmortem blood specimens and as a marker of lurasidone administration in the event lurasidone is degraded to concentrations below detection limits.

Simultaneous quantification of thalidomide, lenalidomide and pomadomide in plasma by LC-MS/MS

ObjectiveTo develop a new method for quantitatively analyzing three immunomodulators (thalidomide, lenalidomide and pomadomide) by liquid chromatography tandem mass spectrometry (LC-MS/MS). MethodsUsing thalidomide-d4 as internal standard, the three analytes were separated on Agilent Zorbax SB-C18(2.1 mm × 100 mm, 3.5 μm, Agilent, USA) column and monitored in multiple reactions monitoring mode in Agilent G6460A triple quadrupole mass spectrometer operating in positive ionization mode. The sample was pretreated by protein precipitation using methanol at 3-fold volume to sample. The mobile phase was comprised of 0.1% formic acid in water (phase A) and acetonitrile (phase B) and was delivered in gradient elution program. The flow rate was 0.3 mL/min, and the injection volume was 5 μL. ResultsThe accuracy and stability of the method are within ±15.0%, and the precision is not >15.0%. The recoveries were 85.04% ∼ 119.07%, and the matrix effect was 73.68% ∼ 116.75%. Specificity, linearity, LLOQ, carry-over and dilution were all in line with the requirements of pharmacopeia and guidelines. The peak concentrations of thalidomide, lenalidomide shows huge inter-individual differences. ConclusionsThis newly developed method was sensitive, simple, and robust and can be used in therapeutic drug monitoring of three immunomodulators in multiple myeloma patients.

A sensitive liquid chromatographic-mass spectrometry method for the quantification of vincristine in whole blood collected with volumetric absorptive microsampling

Vincristine is a well-established cytotoxic drug. In paediatric populations blood collection via venipuncture is not always feasible. Volumetric absorptive microsampling (VAMS) is a less invasive method for blood collection. Furthermore, VAMS lacks the haematocrit effect on the recovery known with dried blood spots. Therefore, a liquid chromatography tandem-mass spectrometry method was developed and validated for the quantification of vincristine in whole blood collected with VAMS devices. Sample preparation consisted of solid-liquid extraction with 0.2% formic acid in water and acetonitrile. The final extract was injected on a C18 column (2.0 ×50 mm, 5 µm). Gradient elution was used and quantification was accomplished with a triple quadruple mass spectrometer operating in the positive mode. The validated concentration range was from 1 to 50 ng/mL with an intra- and inter-accuracy and precision of ± 10.3% and ≤ 7.3%, respectively. This method was able to successfully quantify vincristine concentrations in whole blood collected with VAMS from paediatric oncology patients. Vincristine concentrations in whole blood were non-linearly associated with plasma concentrations, which could be described with a saturable binding equilibrium model.

Development and validation of LC–MS/MS method for quantification of protease inhibitor Pepstatin A to monitor its robust clearance in vaccine downstream process.

Pepstatin A reversibly inhibits aspartic acid proteases and minimizes the impact of protease-induced degradation in recombinant protein manufacturing process. Pepstatin A is considered as a process-related impurity and must be characterized and controlled during manufacturing. Herein we describe the development and validation of an LC-MS/MS method for the quantitation of pepstatin A to monitor its robust clearance in vaccine purification process. Analyte extraction from process intermediates was carried out using 10% acetonitrile/water extraction method. Acetyl-pepstatin was used as internal standard (IS). Pepstatin A and IS were resolved on a C18 column using 10 mM ammonium acetate in water and methanol/acetonitrile mobile phase system. A triple quadrupole mass spectrometer operating in the positive electrospray ionization mode with multiple reaction monitoring was used to detect Pepstatin A and IS transitions of m/z 686.5 to 229.3 and 644.5 to 229.3, respectively. The method was validated for specificity, linearity, accuracy, repeatability (precision), intermediate precision, and assay robustness. The assay was linear over the range of calibration standards 0.5–100 ng/mL. The Lower-limit-of-quantification (LLOQ) of the method was 0.50 ng/mL.

Laboratory rotational spectroscopy and astronomical search of ethynyl substituted naphthalene

ABSTRACT The recent interstellar detection of cyanonaphthalenes brings interest in related aromatic molecular species that could be present in similar astronomical environments. In this context, ethynyl derivatives of naphthalene are promising candidates to be observed in the Taurus Molecular Cloud (TMC-1), where cyanonaphthalenes together with cyano- and ethynyl-derivatives of cyclopentadiene and benzene have been detected. To enable the interstellar searches for ethynyl derivatives of naphthalene, their pure rotational spectra need to be investigated in the laboratory. We have observed for the first time the rotational spectra of 1- and 2-ethynylnaphthalene species using a broad-band Fourier-transform microwave spectrometer operating in the 2–8 GHz frequency region. Accurate spectroscopic parameters are derived from the analysis of the experimental spectra, allowing for reliable predictions for astronomical searches. Our searches in TMC-1 for both isomers provide upper limits for the abundances of these species.

Accelerator Mass Spectrometry “Golden Valley”

Information about the resources of the laboratory “AMS Golden Valley” and the state of affairs in accelerator mass spectrometry (AMS) in Russia is presented. The key differences of the AMS method from traditional methods for determining radiocarbon are described, the principle of operation of accelerator mass spectrometers of Russian (unique scientific facility “AMS BINP SB RAS”) and Swiss (MICADAS-28) production is given, and basic information is given about the methods for preparing graphite targets for AMS-analysis.

Are Plants Capable of Pheomelanin Synthesis? Gas Chromatography/Tandem Mass Spectrometry Characterization of Thermally Degraded Melanin Isolated from Echinacea purpurea

Echinacea is a widely used plant medicine, valued especially for its well-documented ability to stimulate the immune system. It has been suggested that melanin could be one of the bioactive factors responsible for the immunostimulatory properties of the plant. The biological functions of melanin pigments are closely related to their chemical composition and structural features. The aim of this study was to characterize the melanin from Echinacea purpurea based on the analysis of thermal degradation products of the well-purified pigment extracted from the dried herb. The melanin was pyrolyzed, and the resulting products were separated by gas chromatography and identified using a triple quadrupole mass spectrometer operating in full scan and multiple reaction monitoring modes. Three groups of marker products were detected in the melanin pyrolysate: polyphenol derivatives, nitrogen-containing heterocycles devoid of sulfur, and benzothiazines/benzothiazoles. This suggests that E. purpurea produces three structurally different melanin pigments: allomelanin, eumelanin, and pheomelanin, which in turn may affect the biological activity of the herb. Our results provide the first-ever evidence that plants are capable of synthesizing pheomelanin, which until now, has only been described for representatives of the animal and fungal kingdoms.

In vitro degradation of ziprasidone in human whole blood.

A systematic study was performed into the degradation of ziprasidone in simulated postmortem blood. Fifteen potential degradation products not previously reported in the literature were observed. Four resulted from degradation in human blood, whereas the remaining products resulted from reaction with solvents: four from alkaline degradation, four from reaction with acetaldehyde, and three from reaction with acetone. To identify possible degradation products, a liquid chromatograph-diode array detector (LC-DAD) and liquid chromatograph quadrupole-time-of-flight mass spectrometer (LC-QTOF-MS) operating in auto-MS/MS mode were used. It was indicated from red-shifted UV-Vis spectra, accurate mass data, mass fragmentation data, and a deuteration experiment that the site of ziprasidone degradation, in the in vitro blood experiments, was the methylene carbon of the oxindole moiety. The major in vitro blood degradation products were proposed to be E/Z isomers of 3-ethylidene-ziprasidone. Further, another in vitro degradation product in microbially inoculated blood specimens was proposed to be 3-ethyl-ziprasidone. 3-Ethylidene-ziprasidone was hypothesized to form from the reaction of ziprasidone with acetaldehyde derived from the ethanol used to spike ziprasidone into the in vitro blood experiments. Data from two postmortem investigations were available for retrospective reanalysis. Attempts were made to detect degradation products of ziprasidone, but none were found.

Development and application of a liquid chromatography coupled to mass spectrometry method for the simultaneous determination of 23 antineoplastic drugs at trace levels

The goal of this study was to develop a method for the simultaneous quantification of 23 commonly used antineoplastic drugs in a hospital pharmacy, using ultra-high pressure liquid chromatography separation coupled to tandem mass spectrometry detection (UHPLC-MS/MS). The following drugs were investigated: 5-fluorouracil, cytarabine, ganciclovir, gemcitabine, dacarbazine, methotrexate, pemetrexed, busulfan, topotecan, rentitrexed, ifosfamide, cyclophosphamide, etoposide, irinotecan, doxorubicin/epirubicin, vincristine, docetaxel, paclitaxel, daunorubicin, idarubicin, vinblastine, oxaliplatin and carboplatin. The chromatographic separation was performed on a phenyl-hexyl column (2.1 ×100 mm, 1.7 µm) with a gradient elution of methanol and water containing 10 mM ammonium formate adjusted to pH 4.9. All compounds were analyzed in less than 13 min and detected with a triple quadrupole mass spectrometer operating in MRM mode. Limits of detection (LODs) and limits of quantification (LOQs) were comprised between 0.01 and 5 ng.mL−1, and between 0.5 and 5 ng.mL−1, respectively. Accuracies ranged between 117% and 83% at the LOQ, intermediate and upper LOQ concentrations, with relative standard deviations (RSD) inferior to 8%, for all the antineoplastic drugs. Finally, the UHPLC-MS/MS method was successfully applied to the analysis of surface samples to evaluate the chemical contamination by these highly toxic compounds in a chemotherapy preparation unit in a hospital pharmacy with the purpose of monitoring the exposure of health care professionals.

Simple and high sample throughput LC/ESI-MS/MS method for bioequivalence study of prazosin, a drug with risk of orthostatic hypotension

Objective The study aimed to develop a rapid, simple and sensitive LC/ESI-MS/MS method to measure prazosin concentration in human plasma and apply bedside sampling in bioequivalence study of two prazosin tablets to resolve the adverse effect of orthostatic hypotension. Significance The LC/ESI-MS/MS prazosin method was highly sensitive and selective. Bedside sampling reduced the orthostatic hypotension incidence and subject dropout rate. Methods After sample preparation, prazosin and terazosin (IS) were detected on mass spectrometer operating in multiple reaction monitoring mode using positive ionization. Mobile phase flow rate was set at 0.40 mL/min with sample run time of 1.75 min. The bioanalytical method was validated as per EMEA and FDA guidelines. Bedside sampling was performed in bioequivalence study for the first 4 h after dosing. The three primary pharmacokinetic parameters, C max, AUC0- t and AUC0-∞ and 90% confidence interval were determined. Results The small injection volume of 1 μL minimized instrumentation contamination and prolonged the analytical column lifespan. Linearity was obtained between 0.5 and 30.0 ng/mL, with coefficient of determination, r 2 ≥ 0.99. The mean extraction recovery of prazosin and IS was >92%, with precision value (CV, %) ≤ 10.3%. Only two orthostatic hypotension adverse events were reported. The two prazosin formulations were found to be bioequivalent. Conclusion The LC/ESI-MS/MS method has shown robustness and reliability exemplified by the incurred sample re-analysis result. Bedside sampling should be proposed for bioequivalence or pharmacokinetic studies of drugs demonstrating adverse event of orthostatic hypotension.