[Simultaneous determination of 102 synthetic cannabinoids in electronic cigarette oil by liquid chromatography-tandem mass spectrometry].

Synthetic cannabinoids (SCs), which are among the most widely abused new psychoactive substances, are much more potent and have greater efficacy than natural cannabis. SCs can be disguised in various ways and are commonly sold in the form of electronic cigarette oil. SCs belong to a large family with structures consisting of a core with substituents, linker, ring with substituents, and tail. New SCs can be developed by adding substituents, such as halogen, alkyl, and alkoxy groups, to the aromatic ring system or by changing the alkyl chain length. Since the emergence of so-called first-generation SCs, subsequent developments have led to eighth-generation indole/indazole amide-based SCs. As of July 1, 2021, the entire category of SCs was added to the list of controlled substances, but implementation requires urgent improvements in detection technologies. Typically, each method is limited to a few SCs. Owing to the vast number of chemically diverse SCs and their fast update speed, the determination and identification of various types of SCs using a single method is challenging. Therefore, rapid, sensitive, and accurate quantitative methods that includes various types of SCs must be developed to meet the demand for the qualitative and quantitative analysis of new SCs in seized electronic cigarette oil. In this study, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the simultaneous determination of 102 SCs in electronic cigarette oil. The mass spectrometry and liquid-phase conditions influencing SC separation and determination were optimized. Using the external standard method, 102 SCs were successfully identified in electronic cigarette oil. The samples were extracted using methanol. Target analytes were separated on a Shimadzu Shim-pack GIST-HP C18 AQ column (100 mm×2.1 mm, 1.9 µm) at a column temperature of 40 ℃. The mobile phases consisted of (A) 0.1% formic acid aqueous solution and (B) methanol-acetonitrile (1∶1, v/v). The gradient elution conditions were as follows: 0-8 min, 55%A-15%A; 8-15 min, 15%A; 15-16 min, 15%A-55%A; 16-18 min, 55%A. The flow rate was 0.4 mL/min and the injection volume was 1 µL. Operating in the multiple reaction monitoring mode, the 102 SCs were identified within 18 min. Each SC exhibited a good linear relationship in the range of 1-100.0 µg/L with a correlation coefficient (r)≥0.9915. The limits of detection were 0.01-0.30 µg/L and the limits of quantification were 0.04-0.99 µg/L, which meet the requirements for analyzing SCs in actual samples. Precision was determined using standard solutions with 2, 10, and 50 µg/L of the SCs. The precisions (n=6) were 0.3%-6.0%. The recoveries of the 102 SCs, as evaluated by spiking electronic cigarette oil at low (2 µg/mL), medium (10 µg/mL), and high (50 µg/mL) levels, were 80.1%-119.8%. Good performance was observed for the analysis of real samples. The developed method is accurate, rapid, sensitive, and effective for the determination of the 102 SCs in electronic cigarette oil, satisfying the requirements for practical qualitative and quantitative analysis.

AmTPS6 promotes trehalose biosynthesis to enhance the Cd tolerance in mangrove Avicennia marina.

Cadmium (Cd) pollution poses a significant ecological risk to mangrove ecosystems. Trehalose has excellent potential to mitigate the adverse effects of heavy metals. Unfortunately, the mechanisms related to trehalose-mediated heavy metal tolerance in plants remain elusive. In the present study, we firstly found that Cd induced the accumulation of trehalose and the differential expression of trehalose biosynthesis genes in the roots of mangrove plant Avicennia marina. Then, we found that the application of exogenous trehalose could alleviate the negative effects of Cd on A. marina by phenotypic observation. In addition, photosynthetic parameters and cellular ultrastructure analyses demonstrated that exogenous trehalose could improve the photosynthesis and stabilize the chloroplast and nuclear structure of the leaves of A. marina. Besides, exogenous trehalose could inhibit the Cd2+ influx from the root to reduce the Cd2+ content in A. marina. Subsequently, substrate sensitivity assay combined with ion uptake analysis using yeast cells showed that several trehalose biosynthesis genes may have a regulatory function for Cd2+ transport. Finally, we further identified a positive regulatory factor, AmTPS6, which enhances the Cd tolerance in transgenic Arabidopsis thaliana. Taken together, these findings provide new understanding to the mechanism of Cd tolerance in mangrove A. marina at trehalose aspect and a theoretical basis for the conservation of mangroves in coastal wetlands.

Trehalose along with ABA promotes the salt tolerance of Avicennia marina by regulating Na+ transport.

Mangroves grow in tropical/subtropical intertidal habitats with extremely high salt tolerance. Trehalose and trehalose-6-phosphate (T6P) have an alleviating function against abiotic stress. However, the roles of trehalose in the salt tolerance of salt-secreting mangrove Avicennia marina is not documented. Here, we found that trehalose was significantly accumulated in A. marina under salt treatment. Furthermore, exogenous trehalose can enhance salt tolerance by promoting the Na+ efflux from leaf salt gland and root to reduce the Na+ content in root and leaf. Subsequently, eighteen trehalose-6-phosphate synthase (AmTPS) and 11 trehalose-6-phosphate phosphatase (AmTPP) genes were identified from A. marina genome. Abscisic acid (ABA) responsive elements were predicted in AmTPS and AmTPP promoters by cis-acting elements analysis. We further identified AmTPS9A, as an important positive regulator, that increased the salt tolerance of AmTPS9A-overexpressing Arabidopsis thaliana by altering the expressions of ion transport genes and mediating Na+ efflux from the roots of transgenic A. thaliana under NaCl treatments. In addition, we also found that ABA could promote the accumulation of trehalose, and the application of exogenous trehalose significantly promoted the biosynthesis of ABA in both roots and leaves of A. marina. Ultimately, we confirmed that AmABF2 directly binds to the AmTPS9A promoter in vitro and in vivo. Taken together, we speculated that there was a positive feedback loop between trehalose and ABA in regulating the salt tolerance of A. marina. These findings provide new understanding to the salt tolerance of A. marina in adapting to high saline environment at trehalose and ABA aspects.

[Simultaneous determination of five indole/indazole amide-based synthetic cannabinoids in electronic cigarette oil by ultra performance liquid chromatography].

Synthetic cannabinoids (SCs), which are considered some of the most widely abused new psychoactive substances available today, are much more potent than natural cannabis and display greater efficacy. New SCs can be developed by adding substituents such as halogen, alkyl, or alkoxy groups to one of the aromatic ring systems, or by changing the length of the alkyl chain. Following the emergence of the so-called first-generation SCs, further developments have led to eighth-generation indole/indazole amide-based SCs. Given that all SCs were listed as controlled substances on July 1, 2021, the technologies used to detect these substances must be quickly improved. Due to the sheer number of SCs, the chemical diversity and the fast update speed, it is challenging to determine and identify the new SCs. In recent years, several types of indole/indazole amide-based SCs have been seized, but systematic research on these compounds remains limited. Therefore, developing rapid, sensitive, and accurate quantitative methods to determine new SCs are of great importance. Compared with high performance liquid chromatography (HPLC), ultra performance liquid chromatography (UPLC) shows higher resolution, better separation efficiency, and faster analysis speeds; thus, it can meet the demand for the quantitative analysis of indole/indazole amide-based SCs in seized materials. In this study, a UPLC method was developed for the simultaneous determination of five indole/indazole amide-based SCs, including N-(1-amino-3,3-dimethyl-1-oxobutan-2-yl)-1-butyl-1H-indazole-3-carboxamide (ADB-BUTINACA), methyl 2-(1-(4-fluorobutyl)-1H-indole-3-carboxamido)-3,3-dimethylbutanoate (4F-MDMB-BUTICA), N-(1-methoxy-3,3-dimethyl-1-oxobutan-2-yl)-1-(5-fluoropentyl)-1H-indole-3-carboxamide (5F-MDMB-PICA), methyl 3,3-dimethyl-2-(1-(pent-4-en-1-yl)-1H-indazole-3-carboxamido)butanoate (MDMB-4en-PINACA), and N-(adamantan-1-yl)-1-(4-fluorobutyl)-1H-indazole-3-carboxamide (4F-ABUTINACA) in electronic cigarette oil; these SCs have been detected with increasing frequency in seized materials in recent years. The main factors influencing the separation and detection performance of the proposed method, including the mobile phase, elution gradient, column temperature, and detection wavelength, were optimized. The proposed method successfully quantified the five SCs in electronic cigarette oil via the external standard method. The samples were extracted using methanol, and the target analytes were separated on a Waters ACQUITY UPLC CSH C18 column (100 mm×2.1 mm, 1.7 µm) at column temperature of 35 ℃ and flow rate of 0.3 mL/min. The injection volume was 1 µL. The mobile phase consisted of acetonitrile and ultrapure water, and gradient elution was employed. The detection wavelengths were 290 and 302 nm. The five SCs were completely separated within 10 min under optimized conditions and showed good linear relationships between 1-100 mg/L, with correlation coefficients (r2) of up to 0.9999. The limits of detection (LOD) and quantification (LOQ) were 0.2 and 0.6 mg/L, respectively. Precision was determined using standard solutions of the five SCs at mass concentrations of 1, 10, and 100 mg/L. The intra-day precision (n=6) was <1.5%, and the inter-day precision (n=6) was <2.2%. Accuracy was determined by spiking electronic cigarette oil with low (2 mg/L), moderate (10 mg/L), and high (50 mg/L) levels of the five SCs, with six replicates per determination. The recoveries of the five SCs were 95.5%-101.9%, and their relative standard deviations (RSDs, n=6) were 0.2%-1.5%, with accuracies ranging from -4.5% to 1.9%. The proposed method showed good performance when applied to the analysis of real samples. It is accurate, rapid, sensitive, and effective for the determination of five indole/indazole amide-based SCs in electronic cigarette oil. Thus, it satisfies the requirements for practical determination and provides a reference for the determination of SCs with similar structures by UPLC.

The median effective volume of ultrasound-guided thoracic paravertebral nerve block with 0.3% ropivacaine in radical thoracoscopic surgery for lung cancer.

BACKGROUND: Ultrasound-guided needle placement has revolutionized the thoracic paravertebral block technique and can be applied in thoracoscopic surgery. OBJECTIVE: This study investigated the median effective volume (EV50) of an ultrasound-guided single shot of 0.3% ropivacaine used as a thoracic paravertebral nerve block for the radical thoracoscopic resection of lung cancer. METHODS: A total of 27 patients who received a single shot of ultrasound-guided thoracic paravertebral nerve block and underwent radical thoracoscopic resection of lung cancer were enrolled in this study between February 10 and August 13, 2018. All patients were rated as ASA grades I or II. Using ultrasound as a guide, the block needle was gradually pushed through the lateral costotransverse ligaments to the thoracic paravertebral space by the in-plane technique. After confirming the absence of blood or cerebrospinal fluid, 1-2 ml of 0.3% ropivacaine hydrochloride was injected to confirm that the position of the needle was appropriate, and a pre-determined volume of 0.3% ropivacaine hydrochloride was then administered to the patients. Sensory testing by pinprick was performed every 5 minutes for 30 minutes following the thoracic paravertebral block injection to identify the time segments during which the loss of sensation to the pinprick and its blocking effect occurred. RESULTS: All patients completed the study and 14 (51.8%) had a successful block. CONCLUSION: The EV50 of 0.3% ropivacaine was 18.46 ml (95% CI 17.09-19.95 ml) and the EV95 was 20.89 ml.

A Comparison of Puncture and Continuous Pump Analgesia With Two Different Approaches to Thoracic Paravertebral Block for Thoracic Surgery.

BACKGROUND: This study aims to compare the success rate of thoracic paravertebral block (TPVB) and the effect of postoperative analgesia between two approaches. METHODS: A total of 34 patients with American Society of Anesthesiology (ASA) physical status score II-III, undergoing an optional thoracoscopic surgery, were randomly assigned to a parasagittal approach group (group P, n = 17) and a transverse intercostal approach group (group T, n = 17). The catheterization time, success rate of the puncture and catheterization, block plane and effect at the surgical site were compared between two groups. The mean arterial pressure and heart rate were recorded, as well as the cold tactile block plane and numeric rating scale (NRS) at 0.5, 2, 4, 8, 12, 24, and 48 h after surgery. The study was registered at http://www.chictr.org.cn/showproj.aspx?proj=9624 (Registration number: ChiCTR2100054642). RESULTS: The catheterization time in group P was significantly longer than that in group T (P < 0.05). The success rate of catheterization in group P was lower than that in group T, but no statistical significance (P = 0.085). There was no significant difference in the success rate of Puncture and blocking effect of the surgical site at 30 min post-injection between two groups (P > 0.05). There was no significant difference in the cold tactile block plane and NRS scores during coughing between two groups at 0.5, 2, 4, 8, 12, 24, and 48 h postoperatively (P > 0.05). CONCLUSION: This study suggests that there is no significant difference in postoperative block level or pain score during coughing for thoracoscopic surgery between ultrasound-guided parasagittal and transverse intercostal approach, but the parasagittal approach takes longer and has a higher failure rate.

Monitoring neoadjuvant therapy responses in rectal cancer using multimodal nonlinear optical microscopy.

Most patients with rectal cancer have a better prognosis after receiving neoadjuvant therapy because of its remarkable curative effect. However, no device delivers real-time histopathologic information on treatment response to help clinicians tailor individual therapeutic strategies. We assessed the potential of multimodal nonlinear optical microscopy to monitor therapeutic responses, including tumoral and stromal responses. We found that two-photon excited fluorescence imaging can, without labeling, identify colloid response, inflammatory cell infiltration, vascular proliferation, and tumor regression. It can also directly detect rare residual tumor cells, which may be helpful for distinguishing tumor shrinkage from tumor fragmentation. In addition, second harmonic generation imaging shows the ability to monitor three types of fibrotic responses: mature, intermediate, and immature. We also determined nonlinear spectra, collagen density, and collagen orientation indexes to quantitatively analyze the histopathologic changes induced by neoadjuvant therapy in rectal cancer. Our work demonstrates that nonlinear optical microscopy has the potential to become a label-free, real-time, in vivo medical imaging technique and provides the groundwork for further exploration into the application of nonlinear optical microscopy in a clinical setting.

OGtree: a tool for creating genome trees of prokaryotes based on overlapping genes.

OGtree is a web-based tool for constructing genome trees of prokaryotic species based on a measure of combining overlapping-gene content and overlapping-gene order in their whole genomes. The overlapping genes (OGs) are defined as adjacent genes whose coding sequences overlap partially or entirely. In fact, OGs are ubiquitous in microbial genomes and more conserved between species than non-OGs. Based on these properties, it has been suggested that OGs can serve as better phylogenetic characters than non-OGs for reconstructing the evolutionary relationships among microbial genomes. OGtree takes the accession numbers of prokaryotic genomes as its input. It then downloads their complete genomes from the National Centre for Biotechnology Information and identifies OGs in each genome and their orthologous OGs in other genomes. Next, OGtree computes an overlapping-gene distance between each pair of input genomes based on a combination of their OG content and orthologous OG order. Finally, it utilizes distance-based methods of building tree to reconstruct the genome trees of input prokaryotic genomes according to their pairwise OG distance. OGtree is available online at http://bioalgorithm.life.nctu.edu.tw/OGtree/.