Automated mass spectrometric analysis of urinary and plasma serotonin

Serotonin emerges as crucial neurotransmitter and hormone in a growing number of different physiologic processes. Besides extensive serotonin production previously noted in patients with metastatic carcinoid tumors, serotonin now is implicated in liver cell regeneration and bone formation. The aim was to develop a rapid, sensitive, and highly selective automated on-line solid-phase extraction method coupled to high-performance liquid chromatography–tandem mass spectrometry (XLC-MS/MS) to quantify low serotonin concentrations in matrices such as platelet-poor plasma and urine. Fifty microliters plasma or 2.5 μL urine equivalent were pre-purified by automated on-line solid-phase extraction, using weak cation exchange. Chromatography of serotonin and its deuterated internal standard was performed with hydrophilic interaction chromatography. Mass spectrometric detection was operated in multiple reaction monitoring mode using a quadrupole tandem mass spectrometer with positive electrospray ionization. Serotonin concentrations were determined in platelet-poor plasma of metastatic carcinoid patients (n = 23) and healthy controls (n = 22). Urinary reference intervals were set by analyzing 24-h urine collections of 120 healthy subjects. Total run-time was 6 min. Intra- and inter-assay analytical variation were <10%. Linearity in the 0–7300 μmol/L calibration range was excellent (R² > 0.99). Quantification limits were 30 and 0.9 nmol/L in urine and plasma, respectively. Platelet-poor serotonin concentrations in metastatic carcinoid patients were significantly higher than in controls. The urinary reference interval was 10–78 μmol/mol creatinine. Serotonin analysis with sensitive and specific XLC-MS/MS overcomes limitations of conventional HPLC. This enables accurate quantification of serotonin for both routine diagnostic procedures and research in serotonin-related disorders.     

Mechanism of Intramolecular Photostabilization in Self‐Healing Cyanine Fluorophores

Organic fluorophores, which are popular labels for microscopy applications, intrinsically suffer from transient and irreversible excursions to dark‐states. An alternative to adding photostabilizers at high concentrations to the imaging buffer relies on the direct linkage to the fluorophore. However, the working principles of this approach are not yet fully understood. In this contribution, we investigate the mechanism of intramolecular photostabilization in self‐healing cyanines, in which photodamage is automatically repaired. Experimental evidence is provided to demonstrate that a single photostabilizer, that is, the vitamin E derivative Trolox, efficiently heals the cyanine fluorophore Cy5 in the absence of any photostabilizers in solution. A plausible mechanism is that Trolox interacts with the fluorophore through intramolecular quenching of triplet‐related dark‐states, which is a mechanism that appears to be common for both triplet‐state quenchers (cyclooctatetraene) and redox‐active compounds (Trolox, ascorbic acid, methylviologen). Additionally, the influence of solution‐additives, such as cysteamine and procatechuic acid, on the self‐healing process are studied. The results suggest the potential applicability of self‐healing fluorophores in stochastic optical reconstruction microscopy (STORM) with optical super‐resolution. The presented data contributes to an improved understanding of the mechanism involved in intramolecular photostabilization and has high relevance for the future development of self‐healing fluorophores, including their applications in various research fields.     

X-ray spectrometry

Summary X-ray spectrometry came only into use in Europe as a practical analytical tool in the mid fifties. Development of electronic detectors and registration during and after World War 2 opened this possibility. The method was slowly introduced into the C.S.I. program, but in 1965 at C.S.I 12 the technique had reached maturity. Tables and figures shown in the 1965 survey paper are presented again together with modern achievements for sensitivity, accuracy and ease of operation. Recent improvements concerning excitation, dispersion and detection are discussed. The tremendous influence of the modern computer on both qualitative and quantitative analysis is illustrated, making modern XRFS a very accurate technique. Emphasis of future development will be focussed on sample preparation and handling, reduction of background, leading to better signal-to-noise ratios, and simplification of calculating procedures.

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RöntgenspektrometrieVergangenheit, Gegenwart und Zukunft

Zusammenfassung Erst in der Mitte der fünfziger Jahre hat die Röntgenspektrometrie in der Industrie Eingang gefunden als praktische Analysenmethode. Die Entwicklung der elektronischen Detektoren und Registriergeräte während des zweiten Weltkrieges hatte dazu die Möglichkeit eröffnet. Diese Methode wurde allmählich in das Programm des C.S.I. einbezogen. Beim C.S.I. 12 im Jahre 1965 war dieser neue Zweig der Atomspektrometrie schon ziemlich weit fortgeschritten. Deswegen werden Tabellen und Abbildungen, die damals in dem Übersichtsvortrag gezeigt wurden, nochmals präsentiert mit neuzeitlichen Ergänzungen und Verbesserungen: Empfindlichkeit, Genauigkeit und Bedienungsfreundlichkeit.Ergebnisse der neuesten Entwicklungen auf dem Gebiete der Anregung, der spektralen Zerlegung und der Registrierung werden gezeigt und erläutert.Die stürmische Einführung der Groß-, Klein-, und Kleinstrechner hat einen sehr großen Einfluß sowohl auf die qualitative wie auch auf die quantitative Analyse ausgeübt, und zwar in einem Maße, daß die XRFS heute als die genaueste Analysenmethode betrachtet werden kann. Weitere Fortschritte können erwartet werden für Probenvorbereitung und Behandlung, Erniedrigung des Untergrundes sowie Berechnung der chemischen Konzentration.

     

Recent Emission Channeling Studies in Wide Band Gap Semiconductors

We present results of recent emission channeling experiments on the lattice location of implanted Fe and rare earths in wurtzite GaN and ZnO. In both cases the majority of implanted atoms are found on substitutional cation sites. The root mean square displacements from the ideal substitutional Ga and Zn sites are given and the stability of the Fe and rare earth lattice location against thermal annealing is discussed.     

A new simplicial cover technique in constrained global optimization

A simplicial branch and bound-outer approximation technique for solving nonseparable, nonlinearly constrained concave minimization problems is proposed which uses a new simplicial cover rather than classical simplicial partitions. Some geometric properties and convergence results are demonstrated. A report on numerical aspects and experiments is given which shows that the most promising variant of the cover technique can be expected to be more efficient than comparable previous simplicial procedures.     

Fracture data and stress-strain behavior of rocks in triaxial compression

Test results are reported for a recently completed experimental research program on rocks subjected to triaxial compression. Sandstone, marble, granite and shale specimens were tested at confining pressures as high as 90,000 psi corresponding to mean stresses of up to 143,000 psi. Recognizing that the largest potential experimental error in such tests results from making strain and load measurements external to the vessel, special load and strain-measuring devices were designed and fabricated for use inside the pressure vessel. The specimens were carefully machined cylinders with length-to-diameter ratios of two and with diameters ranging from 4/16 in. to 1 in. The confining pressure was held constant during each run, but varied from 0 to 90,000 psi over the tests. Results are reported in the form of:

1. Stress-strain curves for individual specimens
2. Maximum shear stress at fracture vs. mean-stress curves for each rock type tested
3. Tabulation of results for 59 specimens

A number of tests were run on granite specimens which had been previously fractured. Results from these tests showed good agreement with tests on intact granite, providing the confining pressure was above 30,000 psi.     

A sensitive and rapid bridge for the study of magnetic susceptibilities at frequencies from 200 to 106 Hz

Summary A new piece of equipment is described for measuring / ₀ and / ₀ as a function of field (0 to 4250 Oe), temperature (1.2°K to room temperature) and frequency (200 Hz to 1 MHz). It is about ten times more sensitive than the Hartshorn bridge used in Leiden²⁾ and it is more convenient to operate as the measuring procedure is automatic after initial adjustments have been made. The main component is a bridge circuit of four inductors, built as closely similar to each other as possible. The output from the bridge goes to two phase sensitive detectors which monitor the two outputs, one inphase, one /2 out of phase, of the bridge. A heterodyne system is used where the input signal to the bridge is obtained by mixing the output from a variable frequency oscillator with that from a 1.5 MHz oscillator and taking the difference frequency, which is phase locked to a master oscillator. The output from the bridge, after preamplification, is mixed with a second output from the variable oscillator and the difference taken again. This gives a 1.5 MHz signal modulated by the magnetic effects in the bridge which is used in the two phase-sensitive detectors. Their output is recorded on an x–y writer. The bridge needs only be balanced to an output of about 50 mV because the sample is moved between two coils and the difference voltage is measured. The use of the same equipment to measure relaxation times longer than 100 ms is also described.Communication No. 349a from The Kamerlingh Onnes Laboratorium, Leiden, The Netherlands     

Determining equilibrium constants for dimerization reactions from molecular dynamics simulations

With today's available computer power, free energy calculations from equilibrium molecular dynamics simulations "via counting" become feasible for an increasing number of reactions. An example is the dimerization reaction of transmembrane alpha-helices. If an extended simulation of the two helices covers sufficiently many dimerization and dissociation events, their binding free energy is readily derived from the fraction of time during which the two helices are observed in dimeric form. Exactly how the correct value for the free energy is to be calculated, however, is unclear, and indeed several different and contradictory approaches have been used. In particular, results obtained via Boltzmann statistics differ from those determined via the law of mass action. Here, we develop a theory that resolves this discrepancy. We show that for simulation systems containing two molecules, the dimerization free energy is given by a formula of the form ΔG ∝ ln(P(1) /P(0) ). Our theory is also applicable to high concentrations that typically have to be used in molecular dynamics simulations to keep the simulation system small, where the textbook dilute approximations fail. It also covers simulations with an arbitrary number of monomers and dimers and provides rigorous error estimates. Comparison with test simulations of a simple Lennard Jones system with various particle numbers as well as with reference free energy values obtained from radial distribution functions show full agreement for both binding free energies and dimerization statistics.     

Synthesis, crystal structure and magnetic characterization of metal(II) coordination polymers based on 2-carboxyethylphosphonic acid and 1,10-phenanthroline (metal=Cu, Co, Cd)

Three non-isostructural metal(II) coordination polymers (metal=copper, cobalt, cadmium) were synthesized under the same mild hydrothermal conditions (T=408 K) by mixture of the corresponding metal acetate with 2-carboxyethylphosphonic acid and 1,10-phenanthroline (1:1:1 M ratio) and their structures were determined by single-crystal X-ray diffraction. Cu₂(HO₃PCH₂CH₂COO)₂(C₁₂H₈N₂)₂(H₂O)₂ and Cd₂(HO₃PCH₂CH₂COO)₂(C₁₂H₈N₂)₂ are triclinic (space group P-1) with a=7.908(5) Å, b=10.373(5) Å, c=11.515(5) Å, α=111.683(5)°, β=95.801(5)°, γ=110.212(5)° (T=120 K), and a=8.162(5) Å, b=9.500(5) Å, c=11.148(5) Å, α=102.623(5)°, β=98.607(5)°, γ=113.004(5)° (T=293 K), respectively. In contrast, [Co₂(HO₃PCH₂CH₂COO)₂(C₁₂H₈N₂)₂(μ-OH₂)](H₂O) is orthorhombic (space group Pbcn) with a=21.1057(2) Å, b=9.8231(1) Å, c=15.4251(1) Å (T=120 K). For these three compounds, structural features, including H-bond network and the π-π stacking interactions, and thermal stability are reported and discussed. None of the materials present a long-range magnetic order in the range of temperatures investigated from 300 K down to 1.8 K.