Additional file 1 of Changes in the physiological activity of parenchyma cells in Dalbergia odorifera xylem and its relationship with heartwood formation

Supplementary Material

Quality Similarity between Induced Agarwood by Fungus and Wild Agarwood

To prevent the exploitation of wild agarwood, the development of artificial agarwood through fungal inoculation is a promising method, but finding species that produce efficient high-quality agarwood remains difficult. In this study, a fungal inducer was prepared using wild agarwood containing fungi and high-throughput sequencing was performed to determine its species makeup. Subsequently, it was used to inoculate Aquilaria sinensis(Lour.) Spreng. The induced agarwood (IA), wild agarwood (WA), and nonresinous whitewood (WW) were analyzed for the extract content. In addition, liquid and gas chromatography–mass spectrometry was used to determine the chemical composition of the samples. The results were used to evaluate the quality of the IA. Mortierella humilisLinnem. ex W.Gams, Oidiodendron maius(Barron), and Tolypocladium album(W. Gams) Quandt, Kepler, and Spatafora were the fungal inducers that were discovered to produce agarwood. The extracts from the IA and WA contained 64 and 69 2-(2-phenylethyl)chromones, respectively, while there were none in the WW. Furthermore, 20 (relative content 36.19%) and 27 (relative content 54.92%) sesquiterpenes were identified in the essential oils of the IA and WA, respectively, and none were identified in the WW. The fungal inducer that was prepared from the WA effectively improves the quality of the agarwood, which is extremely similar to that of the WA

Additional file 1 of The DNA barcode identification of Dalbergia odorifera T. Chen and Dalbergia tonkinensis Prain

Supplementary Material

Gas-Phase Thermochemical Properties of Pyrimidine Nucleobases

The gas-phase acidity and proton affinity of thymine, cytosine, and 1-methyl cytosine have been examined using both theoretical (B3LYP/6-31+G*) and experimental (bracketing, Cooks kinetic) methods. This paper represents a comprehensive examination of multiple acidic sites of thymine and cytosine and of the acidity and proton affinity of thymine, cytosine, and 1-methyl cytosine. Thymine exists as the most stable “canonical” tautomer in the gas phase, with a ΔHacid of 335 ± 4 kcal mol−1 (ΔGacid = 328 ± 4 kcal mol−1) for the more acidic N1−H. The acidity of the less acidic N3−H site has not, heretofore, been measured; we bracket a ΔHacid value of 346 ± 3 kcal mol−1 (ΔGacid = 339 ± 3 kcal mol−1). The proton affinity (PA = ΔH) of thymine is measured to be 211 ± 3 kcal mol−1 (GB = ΔG = 203 ± 3 kcal mol−1). Cytosine is known to have several stable tautomers in the gas phase in contrast to in solution, where the canonical tautomer predominates. Using bracketing methods in an FTMS, we measure a ΔHacid for the more acidic site of 342 ± 3 kcal mol−1 (ΔGacid = 335 ± 3 kcal mol−1). The ΔHacid of the less acidic site, previously unknown, is 352 ± 4 kcal mol−1 (345 ± 4 kcal mol−1). The proton affinity is 228 ± 3 kcal mol−1 (GB = 220 ± 3 kcal mol−1). Comparison of these values to calculations indicates that we most likely have a mixture of the canonical tautomer and two enol tautomers and possibly an imine tautomer under our conditions in the gas phase. We also measure the acidity and proton affinity of cytosine using the extended Cooks kinetic method. We form the proton-bound dimers via electrospray of an aqueous solution, which favors cytosine in the canonical form. The acidity of cytosine using this method is ΔHacid = 343 ± 3 kcal mol−1, PA = 227 ± 3 kcal mol−1. We also examined 1-methyl cytosine, which has fewer accessible tautomers than cytosine. We measure a ΔHacid of 349 ± 3 kcal mol−1 (ΔGacid = 342 ± 3 kcal mol−1) and a PA of 230 ± 3 kcal mol−1 (GB = 223 ± 3 kcal mol−1). Our ultimate goal is to understand the intrinsic reactivity of nucleobases; gas-phase acidic and basic properties are of interest for chemical reasons and also possibly for biological purposes because biological media can be quite nonpolar

LC–MS/MS Bioanalysis of Radioligand Therapeutic Drug Candidate for Preclinical Toxicokinetic Assessment

Radioligand therapy (RLT) has gained significant momentum in recent years in the diagnosis, treatment, and monitoring of cancers. In preclinical development, the safety profile of RLT drug candidate(s) is investigated at relatively low dose levels using the cold (non-radioactive, e.g., 175Lu) ligand as a surrogate of the hot (radioactive, e.g., 177Lu) one in the “ligand-linker-chelator” complex. The formulation of the test article used in preclinical safety studies contains a mixture of free ligand (i.e., ligand-linker-chelator without metal) and cold ligand (i.e., ligand-linker-chelator with non-radioactive metal) in a similar molar ratio as seen under the manufacturing conditions for the RLT drug for clinical use, where only a fraction of free ligand molecules chelate the radioactive metal to form a hot ligand. In this very first report of LC–MS/MS bioanalysis of RLT molecules in support of a regulated preclinical safety assessment study, a highly selective and sensitive LC–MS/MS bioanalytical method was developed for the simultaneous determination of free ligand (NVS001) and cold ligand (175Lu-NVS001) in rat and dog plasma. Several unexpected technical challenges in relation to LC–MS/MS of RLT molecules were successfully addressed. The challenges include poor assay sensitivity of the free ligand NVS001, formation of the free ligand (NVS001) with endogenous metal (e.g., potassium), Ga loss from the Ga-chelated internal standard during sample extraction and analysis, “instability” of the analytes at low concentrations, and inconsistent IS response in the extracted plasma samples. The methods were validated according to the current regulatory requirements in a dynamic range of 0.5–250 ng/mL for both the free and cold ligands using a 25 μL sample volume. The validated method was successfully implemented in sample analysis in support of regulated safety studies, with very good results from incurred sample reanalysis. The current LC–MS/MS workflow can be expanded to quantitative analysis of other RLTs in support of preclinical RLT drug development

Differential Mobility Spectrometry Coupled with Multiple Ion Monitoring in Regulated LC-MS/MS Bioanalysis of a Therapeutic Cyclic Peptide in Human Plasma

A differential mobility spectrometry (DMS) in combination with a multiple ion monitoring (MIM) method was developed and validated for quantitative LC-MS/MS bioanalysis of pasireotide (SOM230) in human plasma. Pasireotide, a therapeutic cyclic peptide, exhibits poor collision-induced dissociation (CID) efficiency for multiple reaction monitoring (MRM) detection. Therefore, in an effort to increase the overall sensitivity of the assay, a DMS-MIM approach was explored. By selecting the most abundant doubly charged precursor ion in both the Q1 and Q3 of the mass analyzer in MIM and combining the DMS capability to significantly reduce the high matrix/chemical background noise, this new LC-DMS-MIM method overcomes the sensitivity challenge in the typical MRM method due to poor CID fragmentation of the analyte. Human plasma was spiked with pasireotide with concentrations in the range 0.01–50 ng/mL. Weak cation-exchange solid-phase extraction was employed for sample preparation. The sample extracts were analyzed with a SCIEX QTRAP 6500 system equipped with an ESI source and DMS device. The separation voltage and compensation voltage of the DMS and other parameters of the MS system were optimized to maximize signal responses. The performance of the LC-DMS-MIM assay for quantitative analysis of pasireotide in human plasma was evaluated and compared to those obtained via LC-MRM and LC-MIM without DMS. Overall, the assay sensitivity with DMS-MIM was approximately 5-fold better than that observed in MRM or MIM without DMS. The assay was validated with accuracy (% bias) and precision (% CV) of the QC results at eight concentration levels (0.01, 0.02, 0.05, 0.15, 0.3, 1.5, 15, and 37.5 ng/mL) evaluated ranging from −4.8 to 5.0% bias and 0.7 to 8.6% CV for the intraday and interday runs. The current LC-DMS-MIM workflow can be expanded to quantitative analysis of other molecules that have poor fragmentation efficiency in CID