Developing analytical methods to assure and control the quality of amino acids has long been a challenge for food ingredient, dietary supplement, and pharmaceutical industries due to the high ...polarity and the absence of chromophores in many amino acids; the situation worsens further by the lack of information of impurities that could potentially be introduced during the manufacturing processes. Herein we utilize a four-step strategy including impurity identification, method development, sample analysis, and targeted impurity detection and quantitation to demystify the impurity profiles of amino acids. The effectiveness of the approach is highlighted using histidine as an example. Analysis of histidine manufacturing and degradation processes led to the identification of 12 potential impurities of histidine, including amino acids (arginine, lysine, asparagine, aspartic acid, alanine, and glycine) and non-amino acid impurities (histamine, histidinol, 4-imidazoleacrylic acid, 4-imidazoleacetic acid, β-imidazolelactic acid, and urea). A HILIC method using Poroshell 120 HILIC-Z column (2.1 × 100 mm, 2.7 µm) and a mobile phase system consisting of ammonium formate buffer at pH 3.2 in water and 0.1% formic acid in acetonitrile coupled with a single quadrupole mass spectrometer was developed for the detection and quantitation of the proposed impurities. Evaluation of 11 commercial histidine samples using the developed method revealed distinct impurity profiles, as a fingerprint for each sample; seven of the 12 proposed impurities were detected in histidine samples tested. The developed method was evaluated in terms of specificity, linearity, range, accuracy, precision, and sensitivity (LOQ: 2.5–60.6 ng/mL) for its suitability for compendial applications. Given the high degree of overlap between the proposed and the detected impurities, the approach could be utilized to strengthen USP standards for controlling the quality of histidine. Extension of the strategy to the analysis of other amino acids is currently underway.
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•Implementation of a four-step strategy for impurity profiling of histidine.•Potential impurities of histidine were proposed and verified in actual samples.•A HILIC-MS method for the determination of 12 amino acids and non-amino acids impurities was developed.
We describe a rapid in situ method for detecting agrochemicals on the surface or in the tissue of fruit using a portable mass spectrometer equipped with an ambient ionization source. Two such ...ionization methods, low temperature plasma (LTP) and paper spray (PS), were employed in experiments performed at a local grocery store. LTP was used to detect diphenylamine (DPA) directly from the skin of apples in the store and those treated after harvest with DPA were recognized by MS and MS/MS. These data therefore allowed ready distinction between organic and non-organic apples. DPA was also found within the internal tissue of purchased apples and its distribution was mapped using LTP. Similarly, thiabendazole residues were detected on the skin of treated oranges in a grocery store experiment in which paper spray was performed by wiping the orange surface with a moist commercial lens wipe and then applying a high voltage to ionize the chemicals directly from the wipe. The handheld mass spectrometer used in these measurements is capable of performing several stages of tandem mass spectrometry (up to MS(5)); the compounds on the fruit were identified by their MS/MS fragmentation patterns. Protonated DPA (m/z 170) produced a characteristic MS(2) fragment ion at m/z 92, while thiabendazole was identified by MS(3) using precursor to fragment ion transitions m/z 202 →m/z 175 →m/z 131. These particular examples exemplify the power of in situ analysis of complex samples using ambient ionization and handheld mass spectrometers.
The ICH guidelines recommend reporting thresholds for regular impurities in drug substances at the level of 0.05% or 0.03% (w/w) depending on the maximum daily intake. Therefore, any instrumental ...method of analysis applicable to the impurity analysis should be able to detect and quantify the analytes at those levels. This investigation was designed to verify the suitability of 1H NMR spectroscopy for the detection of impurities, as a first step in the process before attempting quantification. In order to minimize demand on equipment, this study employed a 400 MHz instrument for structural confirmation and signal assignments of choline (1) and O-(2-hydroxyethyl)choline (2), a known impurity. The limit of detection (LOD) of 2 in 10 mg of 1 was established as 0.01% on a 400 MHz instrument and 2% on a 60 MHz (benchtop) NMR spectrometer. Thus, impurities for which quantification is required are readily detected at 400 MHz or above. These results are in contrast to the widespread belief that 1H NMR sensitivity is insufficient for pharmaceutical impurity analysis. The choice of solvent was recognized as a critical parameter for 1H NMR LOD analysis. Furthermore, publicly available NMR raw data (HMDB) proved to be valuable for unveiling the otherwise cryptic information hidden in complex signal patterns via 1H NMR iterative Full Spin Analysis. Finally, the study uncovered the less noticed, yet characteristic, 14N–1H coupling in the -N+(CH3)3 groups, adding strong arguments for the Raw NMR Data Initiative. Collectively, the data prove that the analytical capabilities of high-field NMR easily fulfill the ICH requirements for detection of impurity in the presence of an actual substance of interest which makes it a step closer to achieving regulatory standards.
•The LOD of choline RCA in choline is < 0.01% at 400 Hz, and 2% at 60 MHz.•Demonstrates feasibility of establishing choline purity and identity by (q)NMR.•Entry-level high- and low-field (400/60 MHz) instruments are suitable equipment.•1H NMR can detect choline impurity in the presence of choline at required LOD levels.•QM-based analysis reveals unknown 14N–1H splitting in choline NMR resonance patterns.
A new ambient ionization method--leaf-spray mass spectrometry--is used to detect allergenic urushiols directly from poison ivy (T. radicans) leaves with no sample preparation. These simple ...measurements show all the urushiols previously reported using liquid chromatography mass spectrometry methods. Tandem mass spectrometry analysis of the leaf spray ions confirms the identifications. Enhanced detection of some urushiols was achieved in the negative mode with the addition of chloride anions to the spray solvent.
A planar differential mobility spectrometer (DMS) was coupled to a Mini 10 handheld rectilinear ion trap (RIT) mass spectrometer (MS) (total weight 10 kg), and the performance of the instrument was ...evaluated using illicit drug analysis. Coupling of DMS (which requires a continuous flow of drift gas) with a miniature MS (which operates best using sample introduction via a discontinuous atmospheric pressure interface, DAPI), was achieved with auxiliary pumping using a 5 L/min miniature diaphragm sample pump placed between the two devices. On-line ion mobility filtering showed to be advantageous in reducing the background chemical noise in the analysis of the psychotropic drug diazepam in urine using nanoelectrospray ionization. The combination of a miniature mass spectrometer with simple and rapid gas-phase ion separation by DMS allowed the characteristic fragmentation pattern of diazepam to be distinguished in a simple urine extract at lower limits of detection (50 ng/mL) than that achieved without DMS (200 ng/mL). The additional separation power of DMS facilitated the identification of two drugs of similar molecular weight, morphine (average MW = 285.34) and diazepam (average MW = 284.70), using a miniature mass spectrometer capable of unit resolution. The similarity in the proton affinities of these two compounds resulted in some cross-interference in the MS data due to facile ionization of the neutral form of the compound even when the ionic form had been separated by DMS.
On-line differential ion mobility (DMS) separation enhances analysis of drugs in urine using Mini MS.
Mass spectrometry benefits from a flexible definition which equates it with many aspects of the science of matter in the ionized state. The field continues to expand rapidly, not only to encompass ...larger and more complex molecules through more powerful instruments, but simultaneously towards in-situ measurements made using smaller, more flexible and just-sufficiently-powerful instruments. The senior author has been fortunate to work in mass spectrometry from 1967 to the present and has been involved in a wide range of efforts which have covered analytical, biological, organic, instrumental and physical aspects of the subject. This effort has been made in the company of a remarkable set of colleagues. From this vantage, it is possible to look both backwards and forwards in this prospective and retrospective piece. This presentation involves a personal look at places, people, instruments and concepts engaged in along a path through Mass Spectrometry. The journey goes from Natal, South Africa, via Cambridge, UK, through Kansas and on to Purdue University, in the great state of Indiana. It starts with natural products chemistry and moves to the physical chemistry of fragmentation and energy partitioning on to complex mixture analysis by tandem mass spectrometry and, hence, to the concepts of thermochemical determination by the kinetic method, preparation of materials by ion soft landing, the possible role of amino acid clusters in the origin of homochiral life and the elaboration of a set of ambient ionization methods for chemical analysis performed using samples in their native state. Special attention is given to novel concepts and instrumentation and to the emerging areas of ambient ionization, molecular imaging and miniature mass spectrometers. Personal mass spectrometers appear to be just over the horizon as is the large-scale use of mass spectrometry in field-based analysis, including point-of-care medical diagnostics.
Chlorophenols (CPs) as a mixture of fourteen congeners from mono- to pentachlorophenol were determined using liquid chromatography/electrospray ionization/ion mobility spectrometry (LC/ESI/IMS) to ...describe the response and analytical performance of a mobility spectrometer as a detector for liquid chromatography. The mobility spectrometer was equipped with an interface so that flows from a large bore column could be electrosprayed directly into the drift tube at flow rates up to 500 μL/min without splitting of flow. A linear gradient of the mobile phase from 40% to 90% methanol and 60% to 10% acetic acid (AcOH)–ammonium acetate buffer solution over 40 min with a C18 column provided baseline separations though mobility spectra for CPs were influenced by mobile phase composition. Product ions formed from CPs with ESI included phenoxide anions CPO
−
, AcOH·CPO
−
, CPOH·CPO
−
, and Na
+
·(CPO
−
)
2
and were found to be governed by the drift gas temperature. Ions were identified using LC/ESI/mass spectrometry (MS) and supported by results from computational modeling. Quantitative response was affected by congener structure through the acidities of the OH moiety and by the composition of the mobile phase. Limits of detection ranged from 0.135 mg/L for 2,3,5-trichlorophenol and pentachlorophenol to 2.23 mg/L for 2-chlorophenol; corresponding linear ranges were 20 and 70.
Novel gas-phase ion mobility based devices as well as those that are based on the existing Drift-time Ion Mobility (IMS) and Differential Mobility Spectrometry (DMS) techniques are evaluated for ...their use as gas-phase ion mobility based filters between electrospray ionization (ESI) source and a quadrupole mass spectrometer (MS). Signal/Noise improvements up to 102-fold are achieved with DMS filtering and enhancement of analyte signals from the chemical background allows detecting the peptide ions in a mixture, which are not detectable without DMS pre-filtering due to high levels of background noise in the mass spectra. A dual-shutter IMS drift tube, constructed with resistive glass tubes that provides uniform electric field inside the drift tube and simplifies its construction, is coupled to MS for the first time and shows the separations of protonated caffeine ions from its cluster ions with different mobilities prior to the introduction to MS. Furthermore, two novel Low Mobility Pass Filters (LMPF) with planar and cylindrical designs are studied for their potential to filter ions of lower mobilities from the system by improving the signal-to-noise ratios in the mass spectra. Planar LMPF is constructed from existing DMS drift tube and operates with low amplitude symmetric high frequency waveform in contrast to more complex electronics of DMS where the waveforms with much higher amplitudes and with asymmetric shape are applied. The cylindrical design of LMPF, which offers very simple alternative design for continuous ion filtering, is constructed with two cylindrical conductive tubes separated with gas-tight insulator tube made from teflon. One of the tubes is connected to ground potential and connected to the MS, while the other tube is applied a negative DC voltage to deflect the ions that are produced by ESI operating in the positive mode. This filter also demonstrates a low mobility cut-off by removing solvent ions, and thus, simplifying the ion population in the ESI plume for the measurement with mass spectrometry. Moreover, issues that are related to the operation of IMS drift tubes and have not been addressed throughout the history of IMS, such as the impact properties of an ion shutter to the performance of IMS and the failure of utilizing IMS as stand-alone detectors for liquid chromatographs (LC) at high flow rates are studied. For the studies with ion shutters the mutual effects of electric fields of the ion shutter and the drift tube and their influence to the sensitivity of IMS drift tubes are examined. It is found that the transfer efficiency of ions through the ion shutter is dependent on their mobilities suggesting that the sensitivity of mobility drift tubes equipped with traditional ion shutters is governed by the individual ion mobilities. These experimental findings are supported and elucidated with a computer-modeling software. An IMS drift tube that operates at ambient pressure and equipped with desolvation region and a liquid drain interface to accept the ESI generated aerosol-ions from LC with high flow rate effluent is evaluated to understand the reasons why IMS drift tubes at ambient pressure have failed as stand-alone detectors for LC. Several instrumental parameters, such as the temperature of the drift gas, the composition of aqueous-organic solvent mixture, and the liquid flow rate are studied for their effect to the performance of the drift tube. These studies demonstrate that IMS drift tubes must be maintained at high temperatures to avoid ion-neutral clustering processes that may complicate the data to interpret. Moreover, an increase of water concentration in aqueous buffer-methanol solvent mixture is found to deteriorate the performance of the IMS drift tubes and are attributed as another reason for the failure of IMS to be used as a detector for LC.
Crystal engineering of gossypol using pyrazine as a complexing agent led to the discovery of new families of crystalline inclusion complexes on the basis of a mixed “gossypol–pyrazine” host. ...Depending on the H-bonding ability of the solvent guest molecules, different types of clathrates were formed. In these clathrates, pyrazine molecules incorporate gossypol molecules to relatively robust mixed host matrix through symmetrical H-bonds of the type O–H⋯N.
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