Quantifying water stress of safflower (Carthamus tinctorius L.) cultivars by crop water stress index under different irrigation regimes.

Infrared thermometry allows evaluating water status of the crop by measuring crop water stress index (CWSI), without the need of physical contact to leaves. In order to quantify water stress by CWSI and finding the best irrigation regime a two-year field experiment was conducted in safflower during 2017 and 2018 growing seasons at Darab, Fars province, Iran. Two safflower cultivars (Goldasht and Local Isfahan) and four irrigation regimes consisted of well-watered [Irrigation based on 100% field capacity (FC)], mild (75% FC), severe (50% FC), and most severe (25% FC) water stress were arranged as split plot according to randomized completely block design with four replicates. The relationship between vapor pressure deficit (VPD) and canopy-air temperature differences (Tc-Ta) was plotted under upper (fully stressed) and lower baselines (non-stressed) equations. In two cultivars, by VPD increment, the distance between upper and lower base lines increased. In Goldasht, the upper baseline (Tc-Ta)ul, was 7.8 °C in 2017 and 8.9 °C in 2018. From April to July when air warmed, Tc-Ta differential was increased up to July and the highest seasonal CWSI (0.72-0.77) were obtained in Local Isfahan under most severe water stress. In 2017, under water stress, the highest relative water content (RWC; 55%), color quality (6-7) and water use efficiency (WUE; 2.69 g m-2 mm-1) was observed in Goldasht under mild water stress which was more than 2018 and Local Isfahan. It might be attributed the more tolerance of Goldasht to water stress and lower air temperature and evaporation in the first year. CWSI with total water consumed (R2 = 0.88∗∗), RWC (R2 = 0.87∗∗), color quality (R2 = 0.75∗) and seed yield (R2 = 0.83∗∗) related, negatively. Overall, a mild water stress (75% FC) with 0.28-0.33 seasonal CWSI had higher RWC, color quality, WUE, with an acceptable yield, which could be the best irrigation regime under water deficit conditions for safflower.

An improved liquid chromatography tandem mass spectrometry (LC-MS/MS) method for quantification of dexmedetomidine concentrations in samples of human plasma.

Dexmedetomidine (DMET) is a sedative, analgesic and anxiolytic with minimum adverse respiratory effects. An LC-MS/MS bioanalytical method has been developed and validated to accurately measure DMET concentrations in samples of human plasma. The method overcomes difficulties in the extraction and quantification of DMET due to the fact that it binds strongly to glass and plastic tubes, as well as solid phase extraction (SPE) cartridges. Human plasma (50 µL) was mixed with the internal standard (IS) (DMET-d4) solution (100 µL) and 0.1% formic acid (50 µL) and extracted using Oasis HLB 1 CC (30 mg) solid phase extraction (SPE) cartridges (Waters®). The glass tubes were coated with bovine serum albumin (BSA) 0.5% (20 µL) before eluting DMET and the IS. After evaporation under nitrogen at room temperature, the analytes were reconstituted in 20% acetonitrile in 0.1% formic acid in water and transferred to silanized glass vials. An electrospray ionisation (ESI) mass spectrometry method in positive mode was created and the precursor/product transitions (m/z) were 201.1 → 95.0 (DMET) and 204.9 → 99.0 (IS). The method was robust and fully validated based on the 2012 EMEA guideline for bioanalytical method validation in the concentration range of 0.5-20 ng/mL. Using this assay, we showed that DMET binds strongly to Extracorporeal Membrane Oxygenation (ECMO) circuits, consistent with expectations for small lipophilic compounds.

High-throughput assay for quantification of the plasma concentrations of thiopental using automated solid phase extraction (SPE) directly coupled to LC-MS/MS instrumentation.

Most previous assays for thiopental are time-consuming due to laborious sample extraction steps prior to analysis using gas chromatography or high pressure liquid chromatography. Here, we describe the first high-throughput liquid chromatography - tandem mass spectrometry (LC-MS/MS) method for quantification of thiopental concentrations in samples of human plasma. Robotic on-line solid phase extraction (SPE) was used to elute the analytes of interest from samples of human plasma (50µL) loaded onto C18 SPE cartridges to which were added aliquots (50µL) of internal standard solution (thiopental-d5 100ng/mL) and 0.5% formic acid in water (100µL). Cartridges were washed using 10% methanol in ammonium acetate buffer (50mM, pH 7) before elution with mobile phase comprising 0.1% formic acid in water and acetonitrile with a flow rate of 0.55mL/min using a 7.2min run time. The analytes were separated on a C18 XTerra® analytical column. Mass spectrometry detection was performed using a QTrap 5500 mass spectrometer (AB Sciex) with negative ionisation. The multiple reaction monitoring (MRM) transitions for thiopental and the internal standard were 241→58, and 246→58, respectively. The calibration curve was linear over a range of 6-600ng/mL. Thiopental was stable in human plasma samples for at least 36h in the autosampler, as well as after three cycles of freeze and thaw, and after 3h storage at room temperature. The absolute recovery and matrix effect were 102% and 6.9%, respectively, and the within-run and between-run precision and accuracy were ≤15%. Our method is fully-validated and satisfies the requirements of the 2012 European Medicines Agency (EMEA) guideline for Bioanalytical Method Validation.

High-throughput assay for simultaneous quantification of the plasma concentrations of morphine, fentanyl, midazolam and their major metabolites using automated SPE coupled to LC-MS/MS.

A rapid LC-MS/MS assay method for simultaneous quantification of morphine, fentanyl, midazolam and their major metabolites: morphine-3-ß-D-glucuronide (M3G), morphine-6-ß-D-glucuronide (M6G), norfentanyl, 1'-hydroxymidazolam (1-OH-MDZ) and 4-hydroxymidazolam (4-OH-MDZ) in samples of human plasma has been developed and validated. Robotic on-line solid phase extraction (SPE) instrumentation was used to elute the eight analytes of interest from polymeric SPE cartridges to which had been added aliquots (150 µL) of human plasma and aliquots (150 µL) of a mixture of two internal standards, viz. morphine-d3 (200 ng/mL) and 1'-hydroxymidazolam-d5 (50 ng/mL) in 50 mM ammonium acetate buffer (pH 9.25). Cartridges were washed using 10% methanol in ammonium acetate buffer, pH 9.25 (1 mL, 2 mL/min) before elution with mobile phase comprising 0.1% formic acid in water (A) and acetonitrile (B) with a flow rate of 0.6 mL/min using an 11.5 min run time. The analytes were separated on a C18 X-Terra® analytical column. The linear concentration ranges were 0.5-100 ng/mL for fentanyl, norfentanyl and midazolam; 1-200 ng/mL for 4-hydroxymidazolam, 2.5-500 ng/mL for 1'-hydroxymidazolam and 3.5-700 ng/mL for morphine, M3G, and M6G. The method showed acceptable within-run and between-run precision (relative standard deviation (RSD) and accuracy <20%) for quality control (QC) samples spiked at concentrations of 80% and 50% of the ULOQ, 3 times higher than the LLOQ, and also at the LLOQ. Furthermore, analytes were stable in samples (after mixing with internal standard) for at least 48 h in the autosampler (except for 4-hydroxymidazolam which decreased by 22% after 24 h), 5 h at room temperature and after three cycles of freeze and thaw. No autosampler carry-over was observed and the absolute recovery (the area ratio of analyte in plasma relative to that in ammonium acetate buffer 50 mM, pH 9.25) was in the range 40% (midazolam) to 110% (morphine). The assay was applied successfully to the measurement of the analytes of interest in plasma samples from patients on extracorporeal membrane oxygenation (ECMO).