Liquid-liquid Extraction Procedure Research Articles

An analytical method using salting-out assisted liquid-liquid extraction to quantify ceftriaxone from micro volumes of human serum.

Ceftriaxone (CTRX) is a commonly used cephalosporin antibiotic. It is suggested that monitoring plasma/serum concentrations is helpful for its safe use. This study aimed to develop and validate an analytical method for measuring CTRX concentrations in human serum according to International Conference on Harmonization guideline M10. Ten microliters of serum sample was purified using a salting-out assisted liquid-liquid extraction procedure with magnesium sulfate. The upper layer was then diluted threefold and analyzed using a liquid chromatography-tandem mass spectrometry-based method with a total run time of 12 min. The linear calibration curve was obtained over the concentration range 5-500 μg/ml. The within-run accuracy varied from 0.2 to 6.5%, and the precision was ≤8.0%. The between-run accuracy and precision ranged from 0.7% to 5.6% and ≤6.4%, respectively. Significant carryover was resolved by injecting four blanks after high-concentration CTRX samples. The recovery rates from spiked serum at low and high concentrations were 44.4 and 43.4%, respectively. Other factors, including selectivity, matrix effects, stability, dilution integrity and reinjection reproducibility also met the acceptance criteria. Serum concentrations in 14 samples obtained from two participants receiving 2 g/day of CTRX were successfully determined using this method.

An analytical method using salting-out assisted liquid-liquid extraction to quantify ceftriaxone from micro volumes of human serum.

Ceftriaxone (CTRX) is a commonly used cephalosporin antibiotic. It is suggested that monitoring plasma/serum concentrations is helpful for its safe use. This study aimed to develop and validate an analytical method for measuring CTRX concentrations in human serum according to International Conference on Harmonization guideline M10. Ten microliters of serum sample was purified using a salting-out assisted liquid-liquid extraction procedure with magnesium sulfate. The upper layer was then diluted threefold and analyzed using a liquid chromatography-tandem mass spectrometry-based method with a total run time of 12 min. The linear calibration curve was obtained over the concentration range 5-500 μg/ml. The within-run accuracy varied from 0.2 to 6.5%, and the precision was ≤8.0%. The between-run accuracy and precision ranged from 0.7% to 5.6% and ≤6.4%, respectively. Significant carryover was resolved by injecting four blanks after high-concentration CTRX samples. The recovery rates from spiked serum at low and high concentrations were 44.4 and 43.4%, respectively. Other factors, including selectivity, matrix effects, stability, dilution integrity and reinjection reproducibility also met the acceptance criteria. Serum concentrations in 14 samples obtained from two participants receiving 2 g/day of CTRX were successfully determined using this method.

Chemical Characterization of Terpene-Based Hydrophobic Eutectic Solvents and Their Application for Pb(II) Complexation during Solvent Extraction Procedure.

Solvents prepared from natural terpenes (menthol and thymol), as H-bond acceptors, and a series of organic acids (chain lengths of 8, 10, and 14 C atoms), as H-bond donors, were characterized and tested as reaction media for liquid-liquid extraction purposes. Due to their high hydrophobicity, they seem to be promising alternatives to conventional (nonpolar and toxic) solvents, since they possess relatively less toxic, less volatile, and consequently, more environmentally friendly characteristics. Assuming that the equilibrium is established between solvent and analyte during a ligandless procedure, it can be concluded that those nonpolar solvents can efficiently extract nonpolar analytes from the aqueous environment. Previous investigations showed a wide range of applications, including their use as solvents in extractions of metal cations, small molecules, and bioactive compounds for food and pharmaceutical applications. In this work, hydrophobic solvents based on natural terpenes, which showed chemical stability and desirable physicochemical and thermal properties, were chosen as potential reaction media in the liquid-liquid extraction (LLE) procedure for Pb(II) removal from aqueous solutions. Low viscosities and high hydrophobicities of prepared solvents were confirmed as desirable properties for their application. Extraction parameters were optimized, and chosen solvents were applied. The results showed satisfactory extraction efficiencies in simple and fast procedures, followed by low solvent consumption. The best results (98%) were obtained by the thymol-based solvent, thymol-decanoic acid (Thy-DecA) 1:1, followed by L-menthol-based solvents: menthol-octanoic acid (Men-OctA) 1:1 with 97% and menthol-decanoic acid (Men-DecA) 1:1 with 94.3% efficiency.

Mass Balance Studies of Robenidine Hydrochloride in the Body of Channel Catfish (Ictalurus punctatus).

This study aims to determine the mass balance of robenidine hydrochloride (ROBH) in the body of Channel catfish (Ictalurus punctatus). ROBH was administered orally at a dose of 20 mg/kg; following drug administration, the water samples were collected at predetermined time points (12, 24, 48, 72, 96, 120, 144, and 168 h), the experimental fish were executed after the water samples were obtained at 168 h, and the tissue samples were collected separately from the bones. The water and tissue samples were analyzed by high-performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS) for concentrations of ROBH and its potential major metabolites, 4-chlorohippuric acid (PCHA) and 4-chlorobenzoic acid (PCBA). The tissue samples were prepared using a modified QuEChERS procedure; the water samples were prepared using a liquid-liquid extraction (LLE) procedure. The results show that the recovery rate of ROBH in fish is very low, less than 2% of the total amount of the drug, and the recovery in water can reach 80.7% of the total amount of the drug. The content of PCBA accounted for 42.4% of the total amount of the drug; the content of ROBH accounted for 38.3% of the total amount of the drug. The content of PCHA accounted for less than 1% of the total amount of the drug. The results show that, after a single administration, ROBH is rapidly metabolized in vivo and excreted in the form of ROBH as well as metabolite PCBA. ROBH and PCBA can be used as the main targets for the metabolism detection of ROBH in Channel catfish.

Discrimination amongst olive oil categories by means of high performance-ion mobility spectrometry: A step forward on food authentication

High-performance-ion mobility spectrometry (HPIMS) has been employed for the first time in the analysis of olive oil samples after a simple liquid-liquid extraction procedure. The HPIMS-based methodology herein developed is user friendly, fast (30 s per analysis) and inexpensive. Moreover, solvent consumption is minimal, thus complying with the principles of green chemistry. The potential of the proposed approach in combination with multivariate analysis has been evaluated for classification of olive oil samples according to their organoleptic quality: extra virgin olive oil (EVOO), virgin olive oil (VOO) and lampante olive oil (LOO). In this work, 115 olive oil samples among the three categories, previously tasted by two Panel Tests, were used as reliable analytical standards to calibrate and validate the chemometric models. The average success rate of classifying samples was 68.9% for validating the ternary model (EVOO, VOO, and LOO). However, the binary model to differentiate between non-defective (EVOO) and defective (VOO and LOO) samples provided a mean classification success rate of 88.3%. In addition, EVOO samples were better classified, with a success rate of ∼90%. The proposed strategy could be used as a valuable tool to support Panel Tests in the industry, allowing for a rapid discrimination between EVOO and non-EVOO samples.

B-163 Development and Validation of a New Quantitative LC-MS/MS Method for Vitamin K1 in Plasma and Serum

Abstract Background Vitamin K1 (phylloquinone) is a fat-soluble cofactor; its deficiency depresses synthesis of plasma prothrombin (factor II) and other factors, causes excessive bleeding, and increases the risk of bone fractures and osteoporosis. Biochemical and metabolic studies on vitamin K1 require both highly sensitive and specific techniques. Analytical challenges to quantitation include the presence of interfering lipids, molecular instability, and extremely low physiological levels. High performance liquid chromatography (HPLC) with fluorescence and electrochemical detection (FLD-ECD) methods used for vitamin K1 involve labor-intensive, liquid-liquid extraction procedures and extremely long run times. Our goal was to develop and validate a high throughput and accurate method for measuring vitamin K1 in plasma and serum using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Methods Plasma or serum specimens were mixed with stable isotope internal standard in an extraction solution containing acetonitrile to release vitamin K1 and precipitate proteins. After centrifugation, the supernatant was then extracted using a HybridSPE® (Supelco) plate to remove the remaining precipitated proteins and phospholipids that may contribute to ion-suppression. Certified standards were used to derive a 6-point calibration curve. Extracts (plasma or serum) and calibrators were analyzed on a SCIEX 6500 + LC-MS/MS with a Thermo TX-4 HTLC multiplexed 4-channel system for increased throughput. Chromatographic separation was achieved on a Biphenyl analytical column. Precision, accuracy, linearity, recovery, and stability were evaluated. Random specimen discards, previously submitted for vitamin K1 testing, were used for a split-sample comparison study vs the current HPLC-FLD-ECD method. Results Compared with the HPLC-FLD-ECD method, the LC-MS/MS method employed a simplified automated hybrid solid phase extraction procedure, used less specimen volume (100 µL vs 700 µL), increased total throughput (4 h vs 32 h per 96-well plate), was more sensitive (limit of quantitation [LOQ] 6.1 pg/mL vs 25.0 pg/mL), and provided greater molecular specificity for vitamin K1 through multiple reaction monitoring (MRM). The intra-assay and total inter-assay precision coefficients of variation (CVW, CVT) in plasma samples (n = 100) were: (6.8%, 5.8%) at 100 pg/mL, (3.5%, 3.0%) at 800 pg/mL, and (3.0%, 3.9%) at 2000 pg/mL. In spiked plasma and serum samples (n = 10), the linearity range was 40–20 000 pg/mL and percent recovery were 98%–107%. Least-squares regression analysis comparing the results for split plasma extracts by LC-MS/MS vs HPLC-FLD-ECD (n = 100) yielded correlation coefficients for vitamin K1: 0.9917, y = 0.927x + 0.827, range 40–7000 pg/mL. Vitamin K1 was stable in plasma and in serum stored in amber plastic screw-capped vials for 8 days at 2–8 °C and 30 days at −20 °C. Conclusion We have developed and validated an LC-MS/MS method that provides rapid, sensitive, precise, and accurate evaluation of vitamin K1 in plasma and serum. Advantages over the existing HPLC-FLD-ECD method include increased throughput and improved sensitivity and specificity. Vitamin K1 results obtained using this method can be used by clinicians to guide clinical intervention to improve or normalize metabolism or prevent the potential adverse consequences associated with inadequate dietary intake.

Geochemical and Geospatial Distribution of Organic Contaminants in the Flood Plain of Ekpetiama, Niger Delta Region of Nigeria

This study investigated the geochemical and geospatial distribution of organic contaminants in the floodplain water and sediments of Ekpetiama in the Niger Delta Region of Nigeria. This study is necessary because there are limited data on the level of organic contamination in this section of the Niger Delta Region of Nigeria. The extinction of planktons in Ekpetiama became a source of concern to the residents. This concern is because this section of the coastal plain provides fisher folks with livelihood. So, there was a need to track the source of contamination in this part of the Niger Delta region. Previous studies have suggested a high level of Total Petroleum Hydrocarbon and Total Hydrocarbon Content as possible sources of reduced dissolved oxygen in similar deltaic terrain. A total of 10 water and 10 sediment samples were collected and analyzed in triplicate at an interval of 100 m in the flood plain. A particle size analyzer was used to perform particle size analyses of air-dried sediments. The American Public Health Association method (APHA) was used to do the chemical analysis of the water samples. Here, a liquid-liquid extraction procedure was performed on sediment samples using 30 mL of Dichloromethane (DCM) as the extracting agent. The results were subjected to statistical validation. The mean grain size ranged from 2.37-4.83, kurtosis (1.94-0.49), and skewness (-0.8-0.71). The contaminant indicators (pH, biochemical oxygen demand, chemical oxygen demand, dissolved oxygen and Total Organic Carbon) point to the presence of organic contamination of the flood plain. The results indicated a total petroleum hydrocarbon range of 0.47-0.87 ppm in water and 0.69-0.96 ppm in sediments and a total hydrocarbon content range of 1.10-2.80 ppm in water and 2.56-3.90 ppm in sediment# samples. The results were above the permitted limits of the World Health Organisation. The source of ecological damage is the abnormal concentrations of organic contaminants in the flood plain. These results significantly caused ecosystem damage and human health effects in the food chain. This study provides information to the National Oil Spill Detection and Response Agency for a cleanup process.

Geochemical and Geospatial Distribution of Organic Contaminants in the Flood Plain of Ekpetiama, Niger Delta Region of Nigeria

This study investigated the geochemical and geospatial distribution of organic contaminants in the floodplain water and sediments of Ekpetiama in the Niger Delta Region of Nigeria. This study is necessary because there are limited data on the level of organic contamination in this section of the Niger Delta Region of Nigeria. The extinction of planktons in Ekpetiama became a source of concern to the residents. This concern is because this section of the coastal plain provides fisher folks with livelihood. So, there was a need to track the source of contamination in this part of the Niger Delta region. Previous studies have suggested a high level of Total Petroleum Hydrocarbon and Total Hydrocarbon Content as possible sources of reduced dissolved oxygen in similar deltaic terrain. A total of 10 water and 10 sediment samples were collected and analyzed in triplicate at an interval of 100 m in the flood plain. A particle size analyzer was used to perform particle size analyses of air-dried sediments. The American Public Health Association method (APHA) was used to do the chemical analysis of the water samples. Here, a liquid-liquid extraction procedure was performed on sediment samples using 30 mL of Dichloromethane (DCM) as the extracting agent. The results were subjected to statistical validation. The mean grain size ranged from 2.37-4.83, kurtosis (1.94-0.49), and skewness (-0.8-0.71). The contaminant indicators (pH, biochemical oxygen demand, chemical oxygen demand, dissolved oxygen and Total Organic Carbon) point to the presence of organic contamination of the flood plain. The results indicated a total petroleum hydrocarbon range of 0.47-0.87 ppm in water and 0.69-0.96 ppm in sediments and a total hydrocarbon content range of 1.10-2.80 ppm in water and 2.56-3.90 ppm in sediment# samples. The results were above the permitted limits of the World Health Organisation. The source of ecological damage is the abnormal concentrations of organic contaminants in the flood plain. These results significantly caused ecosystem damage and human health effects in the food chain. This study provides information to the National Oil Spill Detection and Response Agency for a cleanup process.

Breed-related plasma disposition and faecal excretion of eprinomectin after subcutaneous administration in sheep

Eprinomectin is a member of the avermectin family, exhibiting a broad-spectrum activity as well as a minimal elimination by milk in lactating animals. It is known that pharmacokinetic parameters are fundamental to the rational use of a drug and food safety and these parameters are influenced by different factors. The study aimed to evaluate the plasma dispositions faecal excretion profiles of eprinomectin in Awassi and Cukurova Meat Sheep (CMS) following subcutaneous administration at a dose of 0.2 mg/kg body weight. Heparinized blood and faecal samples were collected at various times between 1 h and 21 days. The plasma and faecal concentrations of eprinomectin were analysed by high-performance liquid chromatography using a fluorescence detector following a liquid-liquid phase extraction procedure. It was observed that eprinomectin concentrations in plasma and faeces were significantly affected by breed. The plasma Tlast, Cmax, AUClast, AUC0-∞, AUMClast, and AUMC0-∞ parameters of eprinomectin were significantly higher (P < 0.014) in Awassi sheep (13.88 ± 1.55 d, 29.81 ± 9.03 ng/mL, 108.05 ± 12.36 ng.d/mL, 116.07 ± 9.35 ng.d/mL, 347.77 ± 40.33 ng.d2/mL, and 486.56 ± 166.25 ng.d2/mL, respectively) compared to CMS (10.00 ± 0.00 d, 19.76 ± 4.51 ng/mL, 64.81 ± 10.54 ng.d/mL, 69.41 ± 10.98 ng.d/mL, 192.00 ± 36.12 ng.d2/mL, and 254.19 ± 52.55 ng.d2/mL, respectively). Similarly, the faecal Cmax, AUClast, and AUC0-∞ parameters of eprinomectin were detected significantly greater in Awassi sheep (3373.27 ± 1499.88 ng/g, 12,428.38 ± 2969.69 ng.d/g, and 12,668.39 ± 3047.32 ng.d/g, respectively) than CMS (2192.57 ± 323.99 ng/g, 9622.78 ± 1356.47 ng.d/g, and 9737.12 ± 1390.92 ng.d/g, respectively). The pharmacokinetic differences of eprinomectin observed between sheep breeds could be attributable to physiological characteristics such as the body fat composition of each breed. In addition, the faecal eprinomectin concentrations were about 110 times higher than those in the plasma of both sheep breeds. In line with the data obtained, eprinomectin found in high concentration in faeces may cause ecotoxicological risk for dung fauna and aquatic organisms having high sensitivity to this compound.

Microwave-assisted conversion of fructose to 5-HMF using carbonaceous acidic catalysts

Recently, the catalytic conversion of fructose to 5-hydroxymethylfurfural (5-HMF) has attracted much interest. The idea of a low-cost, sustainable, energy-efficient method is critical to obtain excellent yield and selectivity for the target product 5-HMF. In this work, microwave (MW) irradiation was chosen as a heating source for the conversion of fructose to 5-HMF with the presence of various amorphous carbons containing Brønsted acid sites as catalysts in dimethyl sulfoxide (DMSO) as a solvent. The catalysts’ morphology, surface functional groups, thermal stability, and compositions of the obtained solids were examined by scanning electron microscope (SEM), elemental mapping, thermogravimetric analysis (TGA), Fourier-transform infrared spectroscopy (FT-IR), energy dispersive X-Ray spectroscopy (EDX), Raman spectroscopy and X-ray diffraction (XRD) techniques. The effect of solvent, heating technique, MW power, time, and catalytic loading on the reaction was studied thoroughly. The highest 5-HMF yield of 79.9% and 5-HMF selectivity of 79.7% were achieved with the optimal condition. 5-HMF was easily separated using a liquid-liquid extraction procedure with high purity. Moreover, the catalyst can be quickly recovered and reused. MW irradiation has proved to selectively and efficiently promote the formation of 5-HMF with high yield in a short reaction time. A mechanism for the formation 5-HMF from fructose was also proposed.

Evaluation of acute oral toxicity, anti-diabetic and antioxidant effects of Aloe vera flowers extract.

Aloe vera (L.) Burm.f. is widely used in various traditional systems of medicine worldwide. Since over 5000 years ago, several cultures have used A. vera extract medicinally for conditions ranging from diabetes to eczema. It has been shown to reduce the symptoms of diabetes by enhancing insulin secretion and protecting pancreatic islets. This research study aimed to investigate the in-vitro antioxidant effect, the acute oral toxicity, and the possible pharmacological in-vivo anti-diabetic activity with histological examination of the pancreas of the standardized deep red A. vera flowers methanolic extracts (AVFME). The liquid-liquid extraction procedure and TLC technique were used to investigate chemical composition. Total phenolics and flavonoids in AVFME were quantified by Folin-Ciocalteu and AlCl3 colorimetric methods, respectively. The present study involved evaluating the in-vitro antioxidant effect of AVFME using ascorbic acid as the reference standard, an acute oral toxicity study by using thirty-six albino rats and different concentrations of AVFME (200mg/kg, 2, 4, 8 and 10g/kg b.w.). Furthermore, the in-vivo anti-diabetic study was performed on alloxan-induced diabetes in rats (120mg/kg, I.P.) and two doses of AVFME (200 and 500mg/kg b.w., orally) were used as compared to glibenclamide (5mg/kg, orally) as a standard hypoglycemic sulfonylurea medication. A histological examination of the pancreas was performed. AVFME resulted in the highest phenolic content of 150.44±4.62mg gallic acid equivalent per gram (GAE/g) along with flavonoid content of 70.38±0.97mg of quercetin equivalent per gram (QE/g). An in-vitro study revealed that the antioxidant effect of AVFME was strong as ascorbic acid. The results of the in-vivo studies showed that the AVFME didn't cause any apparent toxicity signs or death in all groups at different doses which proves the safety of this extract with a wide therapeutic index. The antidiabetic activity of AVFME demonstrated a considerable drop in blood glucose levels as glibenclamide, without severe hypoglycemia or significant weight gain which is considered an advantage of AVFME over glibenclamide use. The histopathological study of pancreatic tissues confirmed the protective effect of AVFME on the pancreatic beta-cells. The extract is proposed to have antidiabetic activity through inhibition of α-amylase, α-glucosidase, and dipeptidyl peptidase IV (DPP-IV). Molecular docking studies were conducted to understand possible molecular interactions with these enzymes. AVFME represents a promising alternative source of active constituents against diabetes mellitus (DM) based on its oral safety, antioxidant, anti-hyperglycemic activities, and pancreatic protective effects. These data revealed the antihyperglycemic activity of AVFME is mediated by pancreatic protective effects while significantly enhancing insulin secretion through increasing functioning beta cells. This suggests that AVFME has the potential as a novel antidiabetic therapy or a dietary supplement for the treatment of type 2 diabetes (T2DM).

LC-MS Method Development and Optimization for Small Drug Analysis in Urine

Both amphetamine and methamphetamine are considered to be illegal chemicals, and hence, the purchase, possession, and use of these drugs is forbidden in many nations. Within the fields of forensic and clinical toxicology, there has been a recent uptick in the detection and quantification of illicit substances within urine samples. Objective: To detect and quantify both drugs in urine samples utilizing caffeine as an internal standard with an optimized liquid-liquid extraction procedure. Methods: An alternative rapid and efficient method of liquid chromatography – electron spray ionization – Tandem mass spectrometry (LC – ESI – TMS) was developed and optimized. The chromatographic separation was carried out using an isocratic high-performance liquid chromatography (HPLC) system, and the eluent that was applied was a mixture of 20% acetonitrile and 80% buffer with a pH of 2.6 that included 10mM ammonium acetate and 0.1% trifluoroacetic acid. The run duration was 9 minutes, and the detection was accomplished at 210 nm with a flow rate of 1 mL/min utilizing triple quadruple MSMS to validate ionic transitions following direct infusion and fragmentation of analytes. Results: An excellent linearity was seen in the calibration curves of amphetamine and methamphetamine in urine samples across the concentration range of 0-10 mg/L, with a regression coefficient of 0.91 and 0.97, respectively, for each of these substances. Conclusions: More compounds are able to be identified in urine as chromatographic techniques, such as high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS), continue to improve in terms of their sensitivity

Focusing on the 5F-MDMB-PICA, 4F-MDMB-BICA synthetic cannabinoids and their primary metabolites in analytical and pharmacological aspects

Nowadays, more and more new synthetic cannabinoids (SCs) appearing on the illicit market present challenges to analytical, forensic, and toxicology experts. For a better understanding of the physiological effect of SCs, the key issue is studying their metabolomic and psychoactive properties.In this study, our validated targeted reversed phase UHPLC-MS/MS method was used for determination of urinary concentration of 5F-MDMB-PICA, 4F-MDMB-BICA, and their primary metabolites. The liquid-liquid extraction procedure was applied for the enrichment of SCs. The pharmacological characterization of investigated SCs were studied by radioligand competition binding and ligand stimulated [35S]GTPγS binding assays.For 5F-MDMB-PICA and 4F-MDMB-BICA, the median urinary concentrations were 0.076 and 0.312 ng/mL. For primary metabolites, the concentration range was 0.029–881.02* ng/mL for 5F-MDMB-PICA-COOH, and 0.396–4579* ng/mL for 4F-MDMB-BICA-COOH. In the polydrug aspect, the 22 urine samples were verified to be abused with 6 illicit drugs. The affinity of the metabolites to CB1R significantly decreased compared to the parent ligands. In the GTPγS functional assay, both 5F-MDMB-PICA and 4F-MDMB-BICA were acting as full agonists, while the metabolites were found as weak inverse agonists. Additionally, the G-protein stimulatory effects of the full agonist 5F-MDMB-PICA and 4F-MDMB-BICA were reduced by metabolites. These results strongly indicate the dose-dependent CB1R-mediated weak inverse agonist effects of the two butanoic acid metabolites.The obtained high concentration of main urinary metabolites of 5F-MDMB-PICA and 4F-MDMB-BICA confirmed the relevance of their routine analysis in forensic and toxicological practices. Based on in vitro binding assays, the metabolites presumably might cause a lower psychoactive effect than parent compounds.

Validated HPLC-UV method for determination of venlafaxine and its metabolite - o-desmethylvenlafaxine in human plasma

This paper describes a simple, rapid and cost-effective HPLC-UV method for the determination of venlafaxine and its active metabolite O-desmethylvenlafaxine in human plasma. Sample preparation is based on a liquid-liquid extraction procedure with a short extraction time. Prazosine hydrochloride was used as the internal standard. The HPLC separation was performed on a Supelcosil LC-CN column, using a mobile phase consisting acetonitrile : 25 mM phosphate buffer at pH 3,5 (15:85 v/v). The calibration curve is linear in the concentrations range of 25-1000 ng/mL, which is suitable for pharmacokinetic studies of venlafaxine hydrochloride following of dosing from 37.5 mg to 375.0 mg per day. The method was fully validated according to the international guidances,

GIS Analysis of Organo-Contaminants and Iron Linked to Groundwater and Sediment at Boreholes in Aluu, Delta Region, Nigeria

The source of high fever and gastrointestinal disorders in humans after groundwater consumption in this part of the delta region in Nigeria is unknown. Spatial data engineered by GIS interpretation of organo-contaminants bound to groundwater and borehole sediment provides baseline data and information on the impact of iron and organo-contaminants on groundwater quality in Aluu. A total of 10 water and sediment samples were collected at a depth of 45 m from 10 boreholes within Aluu and analyzed in triplicate. The choice of 45 m implies the occurrence of sediments bearing groundwater for a deep well. A particle size analyzer was used to perform particle size analyses of the air-dried sediments. The American Public Health Association Method (APHA) was used to perform the chemical analysis of the water samples. Here, a liquid-liquid extraction procedure was conducted on the samples using 30 mL dichloromethane (DCM) as the extraction agent. The results were subjected to statistical validation, spatial data and GIS analysis. The textural characteristics possessed a mean grain size from fine sand (2.03) to medium sand (4.3), poorly sorted of 1.45 to 2.1, skewness of near-symmetrical (0.02), meso-kurtic kurtosis of 0.5 to very platy-kurtic of 2.09. Total petroleum hydrocarbon was 0.033 mg/L to 0.88 mg/L, and total hydrocarbon content and iron were 1.65 mg/L to 3.41 mg/L, and 2.98 mg/L-0.48 mg/L respectively. The results of these contaminants bound to sediments and water were above the acceptable limits of the World Health Organization. The ingress of contaminants into the groundwater was significantly controlled by the characteristics of the borehole sediment.

A Multioutcome, Guided Inquiry-Based Liquid–Liquid Extraction Laboratory for Introductory Organic Chemistry

A multioutcome, guided inquiry-based liquid–liquid extraction (LLE) experiment has been designed for introductory organic chemistry. Multiple outcomes were established by identifying a series of red and blue dyes that are (1) soluble in organic solvent, (2) soluble in aqueous solution, or (3) soluble in either the organic or aqueous layers by tuning the pH of the aqueous solution. This series of dyes led to the development of 10 different dye mixtures that can be separated using LLE. In this experiment, students first evaluate the individual solubilities of three assigned dyes under neutral, acidic, and basic biphasic conditions. The colored nature of these dyes allows students to visually follow their solubilities and thus make connections between solubility and the molecular structure, ionization, and acid/base properties of these compounds. Then, students develop a two-step LLE procedure to separate a three-component mixture of their dyes using the observations from their solubility study, and they perform their separation to verify its effectiveness. Student feedback has shown that the visual nature of this lab permitted >90% of students to prepare a successful separation procedure and that students left this lab feeling more confident about LLE. Further, students felt that this guided inquiry-based experiment allowed them to develop laboratory skills, while also improving their understanding of LLE, enhancing their problem-solving and critical thinking abilities, and increasing their engagement and enjoyment in the lab. The ability to diversify the inquiry problem ensures that all students performing this experiment gain the full benefits of conducting inquiry-based learning.

Highly sensitive MS/MS detection for confident identification of potent novel synthetic opioids and their metabolites

The aim of this presentation is to demonstrate the use of high-resolution mass spectrometry as a unique tool for the untargeted detection and identification of new emerging substances, with little or no method optimization necessary. The presentation will demonstrate how accurate mass instruments are providing extra levels of confidence in analyte identification by reliably obtaining comprehensive MS/MS spectral fragment information that can be used for identification, confirmation, and/or library matching of potent drugs and metabolites present in discarded authentic case samples. NSO compounds were extracted from human whole blood using a liquid-liquid extraction (LLE) procedure. HPLC separation was performed on an ExionLC system using a Phenomenex Kinetex C18 column Mobile phases were ammonium formate and formic acid in methanol and acetonitrile. The flow rate was 0.4 mL/min with a total LC runtime of 15.5 minutes. The injection volume was 10 μL. MS and MS/MS data were collected for each sample using Zeno IDA for optimal sensitivity on the ZenoTOF 7600 system. Data acquisition consisted of a TOF MS scan to collect accurate mass precursor ions from 100 to 700 Da, followed by a TOF MS/MS full scan ranging from 25 to 700 Da to ensure all fragments were captured for identification using a maximum of 16 candidate ions. Data was acquired using SCIEX OS software 2.1. A series of 9 calibrator solutions were prepared by spiking control human whole blood samples with the 6 targeted analytes at final concentrations ranging from 10 pg/mL to 100 ng/mL. The series of calibrator solutions were injected to evaluate the quantitative performance of the system and its ability to accurately measure low level analytes with a high level of precision and accuracy in TOF MS mode. Each calibrator was injected in triplicate. LLOQs of between 10 and 50 pg/ml were observed for all compounds analysed. Accuracies at LLOQ ranged from 86–111% and imprecision was < 15% for all compounds analysed. In addition, the use of specific technologies afforded by the new SCIEX ZenoTOF 7600 system generated an average 9x sensitivity improvement on spectral data providing significantly improved library matching. A comprehensive and highly sensitive method for the screening and identification of potent NSO in human whole blood is described. The significant gains in MS/MS sensitivity on the SCIEX ZenoTOF 7600 system yielded an improvement in confident identifications of low-level analytes through spectral library matching. The observed sensitivity gains resulted in a 9x improvement, on average, in TOF MS/MS sensitivity across the drugs positively identified in the authentic case samples analyzed. This improvement enabled confident identification of key drugs and metabolites at trace levels that were not previously achievable. The MS/MS sensitivity levels afforded by ZenoTOF 7600 system provide a means to monitor low levels of ultra-potent NSO in poly-drug intake scenarios.

Reaching new sensitivity levels for the detection of fentanyl analogs and highly potent novel synthetic opioids in blood

The aim of this presentation is to demonstrate a sensitive quantitative workflow for the analysis of 32 of potent substances in poly-drug, discarded postmortem case samples at low (pg/mL) levels. The presentation will demonstrate that the unparalleled quantification performance of the method enabled accurate detection of potent substances in poly-drug, discarded postmortem case samples at trace levels that were not previously achievable, providing toxicologists a valuable insight into the causation of accidental overdoses. A total of 32 NSO including 17 fentanyl analogs and 15 newly emerging non-fentanyl opioids were selected for this panel. NSO were extracted from the 200 μL of spiked whole blood mixtures using a liquid-liquid extraction (LLE) procedure. Analytes were chromatographically separated using a Phenomenex Kinetex C18 column (50 × 3.0 mm, 2.6 μm, 00B-4462-Y0). Mobile phase A (MPA) and mobile phase B (MPB) were ammonium formate with formic acid and formic acid in methanol and acetonitrile, respectively. The injection volume was 10 μL and the total LC runtime was 15.5 minutes. A single acquisition method consisting of 68 MRM transitions (64 for the NSO and 4 for the internal standards) was created using the Scheduled MRM Algorithm. Control human whole blood samples spiked with the 32 target analytes were prepared at concentrations ranging from 1 pg/mL to 100 ng/mL. Excellent linearity was observed across the concentration ranges analyzed with R2 values greater than 0.99 for all of the NPS in the panel. Reported LLOQ values ranged between 10 to 50 pg/mL for the 32 analytes in the panel. The accuracy and precision of measurements ranged from 80.50–116.64% and from 0.42–17.80%, respectively. The excellent accuracy and precision were observed over the entire concentration range, including at the LLOQ. Overall, the developed method showed excellent reproducibility and linearity, proving the robustness of the developed method and quantitative performance of the SCIEX 7500 System even at low concentration levels for each of the targeted NSO in this study. The applicability of the method to analyze discarded postmortem case samples from subjects suspected of NPS ingestion resulting in accidental overdoses was investigated. For example, the results from the analysis of one of these case samples showed the successful detection of one NSO and its metabolite: isotonitazene and 5-aminoisotonitazene, as well as fentanyl and four of its analogs/metabolites: beta-hydroxy-fentanyl, norfentanyl, 4-ANPP and acetyl fentanyl at concentrations of 1434.33, 7.93, 599.10, 9756.67, 147.69 and 1465.00 pg/mL, respectively. The results demonstrate that the developed method enabled accurate detection of potent substances in poly-drug, authentic case samples at trace levels that were not previously achievable. A robust and sensitive drug screening workflow for the analysis of 32 potent NSO was successfully developed using the SCIEX 7500 System. Overall, the remarkable quantification performance of the SCIEX 7500 System enabled accurate detection of potent NSO at concentrations that were not previously achievable, providing a means to monitor ultra-potent NSO in overdose scenarios. The sensitivity levels afforded by the SCIEX 7500 system provided the ability to detect low levels of NSO in postmortem case samples that would normally go undetected, providing a clearer picture for help in determining the cause of death.

Enhancing high-resolution mass spectrometry performance for NPS analysis with improved sensitivity and characterization

The aim of this presentation is to introduce the instrument features on the ZenoTOF 7600 system that provide qualitative flexibility combined with quantitative power for NPS detection and characterization. The presentation will demonstrate that these new technological advancements on the system can be leveraged to provide more confidence in the quantified amounts of drugs and metabolites detected in discarded authentic case samples which is critical when determining the cause of death following an accidental overdose. Drugs and metabolites were extracted from human whole blood using a liquid-liquid extraction (LLE) procedure. HPLC separation was performed on an ExionLC system using a Phenomenex Kinetex C18 column Mobile phases were ammonium formate and formic acid in methanol and acetonitrile. The flow rate was 0.4 mL/min with a total LC runtime of 15.5 minutes. The injection volume was 10 μL. MS and MS/MS data were collected for each sample using Zeno IDA for optimal sensitivity on the ZenoTOF 7600 system. Fragmentation was performed using both collision-induce dissociation (CID) and electron-activated dissociation (EAD) to compare the generated fragment ions. Data acquisition consisted of a TOF MS scan to collect accurate mass precursor ions from 100 to 700 Da, followed by a TOF MS/MS full scan ranging from 25 to 700 Da to ensure all fragments were captured for identification using a maximum of 16 candidate ions. Data was acquired using SCIEX OS software 2.1. The use of a hybrid collision cell (which offers an alternative fragmentation capabilities) in combination with the Zeno trap technology (which improves MS/MS duty cycle) was leveraged for the characterization of structurally related isomeric species present at low levels in discarded postmortem case samples. The depth of information extracted from the unique fragmentation capabilities of electron-activated dissociation (EAD) enabled differentiation of structurally related isomeric species that were not previously discernable using convention collision-induced dissociation (CID). In addition, the improved MS/MS sensitivity resulted in confident identification of potent novel synthetic opioids and metabolites at concentration levels that were not previously achievable. Overall, the use of the ZenoTOF 7600 system provided a means to characterize and monitor low-levels of ultra-potent NSO in poly-drug intake scenarios. These advancements are shown to support the case of combined opioid drug toxicity leading to death, which offers a clearer picture for help in determining the cause of death. A novel fragmentation technique combined with a highly-sensitive QTOF system for the screening and identification of low-level potent NSO and metabolites in discarded postmortem case samples is described. The depth of information extracted from EAD-based MS/MS spectra combined with the improved MS/MS sensitivity were leveraged for characterization of structurally related isomeric species present at low levels. The results demonstrate that the new technological advancements on the ZenoTOF 7600 provide high levels of confidence in the quantified amounts of drugs and metabolites detected in authentic case samples, which is critical when determining the cause of death following an accidental overdose.

Emergency toxicology analysis – Method development for the straightforward determination of total 70 drugs and metabolites in human blood plasma by LC-MSn

Quantification of 70 emergency toxicology (ET) relevant analytes in human blood plasma using the Bruker Daltonik Toxtyper (TT) system. Rapid and comprehensive qualitative and quantitative methods are crucial in 24/7 ET to support the diagnosis and treatment of acute intoxications. This includes estimating their course and monitoring the effectiveness of possible detoxification. The aim of this study was the simultaneous qualitative and quantitative analysis of 64 drugs and six of their metabolites in human blood plasma by liquid chromatography-ion trap mass spectrometry (LC-MSn, Bruker Daltonik TT). Analytes included compound classes often requested in ET such as analgesics, anticonvulsants, antidepressants, antidiabetics, antiemetics, antihistamines, antihypertensives, benzodiazepines, or neuroleptics. A standard liquid-liquid extraction procedure was used for sample preparation followed by 1:5 dilution of the final extract (Caspar AC. Drug Test Anal 2019;11:102–111). Analyzes were performed using the TT system consisting of a Bruker amazon speed ion trap and a Thermo Fisher Dionex Ultimate 3000 LC system. Plasma concentrations were determined using an electronically stored five-point calibration. Validation was performed according to the European Medicines Agency (EMA) guideline on bioanalytical method validation and the recommendations of the Society of Toxicological and Forensic Chemistry (GTFCh) for ET quantification including selectivity, carry-over, accuracy, precision, and matrix effects. Finally, applicability was tested using 197 patient samples containing 37 different drugs and 5 of their metabolites. The method was selective for all analytes and no relevant carry-over and matrix effect could be observed. A linear and weighted calibration model could be used for all tested analytes. Accuracy and precision recommended for ET could be fulfilled for all tested analytes, except for aripiprazole, diphenhydramine, ketamine, melperone, norpethidine, nortilidine, olanzapine, metformin, pethidine, sertraline, and tilidine. The results after analysis of patient plasma were compared to those obtained by reference LC-tandem MS or gas chromatography-MS methods and data corresponded in case of 190 samples. The concentration levels in the remaining seven samples did not correspond to those determined by one of the reference methods. The present method provides a fast, robust, and reliable blood plasma screening for 70 analytes and an additional quantification of 59 analytes relevant in ET. Due to an electronically stored five-point calibration and a simple “push and print solution”, blood plasma levels of the analytes could be assessed straightforward. Furthermore, the applicability of the method by analyzing ET cases could be demonstrated.