Liquid-liquid Extraction Method Research Articles

Validation of Analytical Method LC MS/MS for Determination Isoniazid in Rats Serum

Determination of isoniazid concentration for therapeutic drug monitoring must produce valid values, so validation of the analytical method of LC-MS/MS needs to be carried out. This research aims to obtain valid method of LCMS/MS in order to determination isoniazid in rats serum. Sample preparation was carried out using the liquid-liquid extraction method using acetonitrile and methanol (2:1). The chromatography system was developed with a C18 column (ACQUITY UPLC® HSS T3 1.8 µm, 2.1 × 100 mm), the mobile phase was 70% acetonitrile with 0.1% formic acid (A), water with 0.1% formic acid (B), as well as methanol (C) with a flow rate of 0.3 mL/min, with gradient elution, detection in ESI+ mode with multiple reaction monitoring (MRM) at the transition 138 m/z → 121 m/z. The research results showed that the method was accurate with a % recovery of 96 - 113% and an RSD of 2.4 ± 0.9%. The precision value was obtained with an average RSD of 3.7 ± 2.4% for intra-day and 4.1 ± 2.3% for inter-day. Linearity showed good results with a correlation coefficient of 0.9956 in tests with 5 different concentrations. The LOQ and LOD values obtained from the linear regression equation were 0.1987 µg/mL and 0.0656 µg/mL respectively. The results showed that the LC-M/MS method was valid for determining isoniazid levels in rat serum

Active Antibacterial Fraction from Ethanolic Extract of Johar Leaves (Cassia siamea Lam.) against Staphylococcus epidermidis and Its Bioautography

Johar leaves (Cassia siamea Lam.) have been traditionally used to treat pruritus due to their antibacterial properties. This study aims to identify the active fraction of an ethanolic extract of Johar leaves against Staphylococcus epidermidis, as well as the active chemical class. The extraction process involved maceration with 96% ethanol. The liquid-liquid extraction method was used to separate the n-hexane, ethyl acetate, and water fractions. The resulting extract and fraction were assessed for antibacterial activity using the well diffusion method. Sample loadings of 4 mg, 2 mg and 1 mg per well were used, with DMSO as a negative control and 6 mg chloramphenicol as a positive control. The test results indicate that the ethyl acetate fraction is more effective than the extract. Specifically, the fraction produced an inhibition zone of 13.5 ± 0.5 mm at a loading of 4 mg/well, while the extract only produced an inhibition zone of 9.3 ± 1.2 mm at the same loading. Additionally, there was a significant difference in the diameter of the inhibitory zone between treatment groups at p 0.05. The TLC results indicate that the ethyl acetate fraction of Johar leaves contains alkaloid, phenolic, flavonoid, and steroid compounds when using silica gel 60 F254 stationary phase and chloroform:ethyl acetate:ethanol (7:2:1 v/v/v) mobile phase. Bioautography results showed that the phenolic and alkaloid compounds at Rf 0.22 and phenolic compound at Rf 0.42 are responsible for the antibacterial activity against Staphylococcus epidermidis.

Low Parts Per Trillion Detection of Iodinated Disinfection Byproducts in Drinking Water and Urine using Vacuum-Assisted Sorbent Extraction and GC-MS/MS.

Disinfection byproducts (DBPs) are ubiquitous environmental contaminants, which are present in virtually all drinking water and linked to detrimental health effects. Iodinated-DBPs are more cytotoxic and genotoxic than chloro- and bromo-DBPs and are formed during disinfection of iodide-containing source water. Liquid-liquid extraction (LLE) paired with gas chromatography (GC)-mass spectrometry (MS) has been the method of choice in the study of low molecular weight iodinated-DBPs; however, this method is laborious and time-consuming and struggles with complex matrices. We developed an environmentally friendly method utilizing headspace solid phase extraction with the application of vacuum to measure six iodinated-trihalomethanes (I-THMs) in drinking water and urine. Vacuum-assisted sorbent extraction (VASE) has the ability to exhaustively and rapidly extract volatile and semivolatile compounds from liquid matrices without the use of solvent. Using VASE with GC-MS/MS provides improved analyte recovery and reduced matrix interference compared to LLE. Additionally, VASE enables extraction of 30 samples simultaneously with minimal sample handling and improved method reproducibility. Using VASE with GC-MS/MS, we achieved quantification limits of 3-4 ng/L. This technique was demonstrated on drinking water from four cities, where five I-THMs were quantified at levels 10-33 times below comparable LLE methods with 10 times lower volumes of sample (10 mL vs 100 mL).

Antioxidant Activity Of Leilem Leaf Extract Fractions Clerodendrum minahassae Teijsm. and Binn. Using The DPPH Method (2,2-Diphenyl-1-Picrylhydrazil)

Clerodendrum minahassae Teijsm. and Binn plant is included in the Clerodendrum genus which contains chemicals such as alkaloids, flavonoids, phenols, tannins and steroids which have an important role in the development of medicine, including as antioxidants, anti-inflammatory, anti-diabetic and antibacterial. This study aims to determine the antioxidant activity of leilem leaf fractions using the DPPH method. Leilem leaves were extracted using the maceration method using 70% ethanol, then partitioned using the liquid-liquid extraction method using three different solvents, namely n-hexane, ethyl acetate and water. The fraction of the partition was then tested for antioxidant activity using the DPPH method and its absorbance was measured using a UV-VIS spectrophotometer with a wavelength of 515 nm. The research results showed that the n-hexane fraction had an IC50 value of 185.704 ppm, the ethyl acetate fraction had an IC50 value of 105.357 ppm and the water fraction had an IC50 value of 549.258 ppm with the IC50 value of the quarcetin comparison being 8.86 ppm. It can be concluded that the highest antioxidant activity is found in the ethyl acetate fraction, which is included in the moderate antioxidant category.

A New Green Solvent: Synthesis and Characterization of Natural-Deep-Eutectic-Solvent (NADES) for Application on Aqueous-Two-Phase System (ATPS) for Extraction of Anthocyanin

Anthocyanins are said to possess antidiabetic, anticancer, anti-inflammatory, antimicrobial, and anti-obesity effects. There is lot of emphasis placed on the improvement of the methods for extracting anthocyanins; one of which being extraction via aqueous two-phase systems (ATPS). Aqueous two-phase system is a liquid-liquid extraction method that has many benefits like easy to scale up with high recovery yield. However, ATPS also has drawbacks as it utilizes organic solvents and low biodegradability ionic liquid. In this context, Natural Deep Eutectic Solvents (NADESs) which are inexpensive, eco-friendly can be employed in aqueous two-phase system. Thus, the purpose of this research is to explore the feasibility of NADES-ATPS in the extraction and purification of anthocyanins. This research will investigate the solubility of anthocyanins and characterize the NADESs in terms of density, viscosity, and polarity to screen out NADESs that are suitable to be used for the extraction of anthocyanins. To do this, 6 different types of NADES with different molar ration, such as 1:1,1:2 and 1:3 of hydrogen bond acceptor (HBA) to hydrogen bond donor (HBD) ratio, were prepared and screened. The NADES was prepared using choline chloride as HBA, and malonic acid, lactic acid, glycerol, ascorbic acid, urea, and citric acid as HBD. The solubility of 10% anthocyanin was found in a range of 0.60×10−2 g/g to 1.81×10−2g/g. The solubility of the anthocyanins in NADES was correlated to the viscosity and polarity of the NADES where it was seen that the solubility has a positive relationship with polarity. The solubility also showed a similar trend to the viscosity however up to critical point, upon which the solvent was too viscous and was unable to be utilized in room temperature. The research concludes that malonic acid-based NADES to be the most suitable to be utilized for the extraction of anthocyanin using ATPS.

Development, validation and application of a liquid chromatography-tandem mass spectrometry method for quantifying lurasidone in dried blood spot samples.

Background: A liquid chromatography-tandem mass spectrometrymethod for quantifying lurasidone in rat dried blood spot (DBS) samples was developed. Method: The analyte was extracted from DBSs using the liquid-liquid extraction method. Chromatographic separation was achieved using a C18, Phenomenex, 150×4.6mm, 3.0μm column. The mobile phase composed of methanol, acetonitrileand water (70:10:20v/v/v) with 0.1% heptafluorobutyric acid performed well in terms of reducing thematrix effect and achieving shorter retention time. Result: The method was validated over a concentration range of 5.0 to 1200.0ng/ml and supported by theevaluation of various validation parameters. Conclusion: This simple, sensitiveand specific method proved to be a viable alternative sampling method with reduced logistics and blood sample storage expenses despite analytical challenges.

Acylhydrazone Functionalized Triphenylamine-Based Fluorescent Probe for Cu2+: Tunable Structures of Conjugated Bridge and Its Practical Application.

Novel fluorescent probes were constructed for the convenient and rapid analysis of Cu2+ ions, taking advantages of the the triphenylamine backbone as chromophore and acylhydrazone group as the Cu2+ recognition site. Especially, probe T2 could act as a dual-channel probe towards Cu2+ through both fluorescent and colorimetric method. Through the fluorescent method, the detection limit of probe T2 was calculated to be as low as 90 nmol/L and there was a good linear relationship between the intensity change and the concentration of Cu2+ ions. By virtue of the two-phase liquid-liquid extraction method, probe T2 could be successfully applied in practical extraction and separation of Cu2+. Furthermore, by applying a "turn-off-turn-on" circle, compound T2 could act as a sensitive probe towards S2- anions through the indirect approach and the detection limit of complex T2-Cu2+ for S2- anion was found to be 110 nmol/L.

Bioanalytical liquid chromatography-tandem mass spectrometry method development and validation for quantification of lurasidone in rat plasma using ion pairing agent.

A bioanalytical method was developed and validated for determining lurasidone (LUR) in rat plasma. The analyte and internal standard were extracted from rat plasma using a liquid-liquid extraction method. The mobile phase consisted of methanol, acetonitrile and water, with an ion pairing agent, 0.1% heptafluorobutyric acid, added to minimise the matrix effect. The detection was achieved using a tandem mass spectrometer (API 2000) in positive ion multiple reaction monitoring mode. All parameters were validated, including selectivity, specificity, carry-over effect, linearity, precision, accuracy, matrix effect, sensitivity and stability. The linearity range was from 5.0 to 1200.0ng/mL with a correlation coefficient of >0.99. The accuracy ranged from 100.00% to 110.22% across the quality control range. The mean absolute recovery from matrix samples for LUR and the internal standard was found to be 68.46% and 67.25%, respectively, and the relative recovery was found to be 73.89% and 77.44%, respectively. This method can determine LUR concentrations in rat plasma samples up to 12 h after oral administration, aiding in LUR pharmacokinetic (PK) investigations in rats. The method's reproducibility on a conventional LC-MS/MS system and a shorter run time of 3.0min make it an appealing bioanalytical method for quantifying LUR in PK studies.

Methods for the relative quantitation of human plasma lipidome using liquid chromatography coupled with mass spectrometry using minimal sample manipulation.

Extraction protocols and liquid chromatography coupled with mass spectrometry (LC-MS) and tandem mass spectrometry (LC-MS/MS) methods for the measurement of four lipid categories, namely glycerophospholipids, glycerolipids, sphingolipids and sterol lipids in human plasma, are described here. Step-by-step instructions are provided for the liquid-liquid extraction methods, including solution preparation and the non-endogenous lipid internal standards used. All instrumental conditions, chromatography columns and solutions required for the LC-MS and LC-MS/MS methods are also provided in detail.

Isolation of Biomolecules from the Leaves of Lecaniodiscus cupanoides (Sapindaceae), a Plant used in Traditional Medicine in Benin

The aim of the present study was to identify the structure of the bioactive molecules in the active ethyl acetate fraction of the hydroethanolic extract of Lecaniodiscus cupanoides (Sapindaceae), a Beninese plant used in the treatment of microbial infections. We prepared the hydroethanolic extract from powdered dried leaves. We fractionated the hydroethanol extract using the liquid-liquid extraction method with solvents of increasing polarity. The active ethyl acetate fraction obtained after bioguided fractionation of the hydroethanol extract on bacterial strains was purified by a series of atmospheric pressure column chromatographic methods coupled with thin layer chromatography. At the end of this purification process, three compounds, including a flavonoid and two fatty acids, were isolated and identified by interpretation of 1H NMR, 13C NMR and mass spectrometry spectra. These were: 2-(3,4-dihydroxyphenyl)-3,5,7-trihydroxy-2,3-dihydrochromen-4-one;(E)-3-(3,4-dihydroxyphenyl)-2-(3-(3,4-dihydroxyphenyl) acryloyloxy) propanoic acid and (E)octadec -9- enoic acid (oleic acid).
 The added value of this work lies in the fact that these three molecules have never before been identified in this plant, and therefore represent a potential avenue for the development of a therapeutic arsenal to combat microbial infections.

The Effect of Using Different Anticoagulant Types for Determination of Esomeprazole Levels in Human Plasma by High-Performance Liquid Chromatography

Esomeprazole is a Proton Pump Inhibitor (PPI) drug formulated in delayed-release tablets, that are included in the mandatory bioequivalence test. In vitro method validation used human plasma from the Indonesian Red Cross that used citrate as an anticoagulant. In the implementation of in vivo study, usually using human plasma used EDTA or heparin as an anticoagulant. This study aims to evaluate the effect of using anticoagulant types that may affect the analysis of esomeprazole in human plasma. Optimum chromatographic conditions used column C18 SunfireTM (5 μm, 250 mm x 4.6 mm); column temperature 40°C; mobile phase acetonitrile - phosphate buffer (40:60% v/v) pH 7.6; 1.0 mL/min flow rate with lansoprazole as an internal standard and wavelength 300 nm (PDA). The extraction was carried out by liquid-liquid extraction method using 500 μl plasma and 5 ml dichloromethane as extraction solvent. The result showed that the concentration range of calibration curve linearity in 5 – 1500 ng/mL. Recovery and broad peak response data of esomeprazole in plasma have significant differences between heparin-EDTA and citrate-EDTA anticoagulants (p<0.05), but there is no significant difference for the stability test. In conclusion, heparin is better than EDTA as an anticoagulant for esomeprazole bioanalysis

Ultrasound-assisted three-phase hollow fiber microextraction-based method for untargeted metabolomics

Sample treatment for untargeted metabolomics is still the bottleneck of the analytical procedure, especially for complex samples like food or biofluids. The main pitfalls are the suppression of minor compounds, low metabolite coverage and the impossibility to detect certain families, combined with the requirement of non-selective approaches for untargeted analysis. Herein, we developed a new analytical extraction method for untargeted metabolomics using a non-selective procedure based on three phase hollow fiber liquid phase microextraction aided by ultrasound (three-phase-UA-HF-LPME) followed by gas chromatography-mass spectrometry. As a proof of concept, the method has been validated for human milk (HM). The analytical method allows extracting organic nitrogenous compounds from HM, which was not possible using the conventional liquid–liquid extraction (LLE) and provided cleaner chromatographic profiles and mass spectra with lower background noise. Moreover, three-phase-UA-HF-LPME enhanced the extraction of a high number of metabolites from 1.65 to 3.27-fold compared to LLE, namely: fatty acids and derivatives (decanoic acid, lauric acid, palmitic acid, tetradecanoic acid, palmitoleic acid, stearic acid, myristic acid, linoleic acid, azelaic acid), lipids and lipid-like molecules (oleamide, monopalmitin), organic acids and derivatives (lactic acid, citric acid), amino acids and peptides (glycine, proline, glutamine). Otherwise, the extraction of carbohydrates and derivatives (urea, phosphoric acid, galactose) decreased the fold changes from 0.15 to 0.25. Results show that our proposed method is attractive owing to its limits of detection from 0.08 to 0.16 mg L−1, recoveries of 99–86 %, enrichment factors up to 123 and fold changes very satisfactory when compared with LLE. The validation by targeted metabolomics as well as the untargeted procedure showed that three-phase-UA-HF-LPME presents better sensitivity, enrichment factors, sample throughput and metabolite coverage than the conventional LLE method considered the gold-standard for untargeted metabolomics. Our results show insights into the possible application of HF for untargeted metabolomics with important analytical advantages in comparison with the LLE.

Development and validation of a deep eutectic solvent-assisted liquid-liquid extraction method for simultaneous quantification of six steroid hormones in serum by liquid chromatography-tandem mass spectrometry

Steroid hormones have been reported to be associated with endocrine system diseases. This paper proposes a novel procedure of deep eutectic solvent (DES)-assisted liquid–liquid extraction (LLE) to extract six steroid hormones (including cortisone, cortisol, androstenedione, testosterone, 17-hydroxyprogesterone, and progesterone) from serum coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS). A total of five types of L-proline, choline chloride, and citric acid-based DESs were tailored; the DES from L-proline and ethylene glycol at a molar ratio of 1:4 with 20 % acetonitrile was selected as the best-fit assisted solvent for the six steroid hormones compared with other DESs. The parameters for extraction by selected DES were optimized using Box-Behnken design (BBD), and the optimal extraction conditions are 200 µL of acetonitrile, 100 µL of the sample, and 80 µL of DES. Under optimum conditions, the method has good linear calibration ranges (between 0.07 ng mL−1 and 600 ng mL−1), correlation coefficients of determination (r2>0.99), and low limits of quantification (between 0.02 and 0.60 ng mL−1). The extraction recoveries were in the range of 81.84–114.43 %, and the intra-day and inter-day relative standard deviations (RSDs) were less than 10 %.In general, the DES-LC-MS/MS method is a simple and environmentally-friendly method, which can be complementary to the presently available methods for determining steroid hormones in serum.

Accurate LC-MS/MS Analysis of Diacylglycerols in Human Plasma with Eliminating Matrix Effect by Phospholipids Using Fluorous Biphasic Extraction.

We developed an accurate method for determining diacylglycerols (DAGs) in human plasma using a fluorous biphasic liquid-liquid extraction method, followed by liquid chromatography with tandem mass spectrometry (LC-MS/MS) analysis. The lipid mixture in the plasma was first extracted with chloroform by using the Bligh-Dyer method. The resulting solution was subjected to fluorous biphasic liquid-liquid extraction to remove phospholipids, which are known to cause matrix effects during the LC-MS/MS analysis. In this method, phospholipids in a lipid mixture solution (nonfluorous solvent) were selectively extracted to tetradecafluorohexane (fluorous solvent) via the specificity of fluorous affinity by forming a complex with a perfluoropolyethercarboxylic acid-lanthanum(III) salt. The remaining DAGs in the nonfluorous solvent could be directly injected into the LC system through the positive electrospray ionization-MS/MS mode. The removal rate of the phospholipids through the fluorous biphasic extraction was more than 99.9%; thus, the matrix-effect-eliminating analysis of DAGs in human plasma with LC-MS/MS was enabled. Furthermore, the applicability of this method and the possibility of using DAGs as biomarkers were evaluated by applying this method to human plasma samples obtained from major depressive disorder as a related disease.

Rapid detection of 10 benzodiazepines and metabolites in blood and urine using DART-MS/MS.

Benzodiazepines are essential screening targets for common sleeping and sedative drugs used in forensic toxicology. Direct analysis in real-time tandem mass spectrometry was used to rapidly identify 10 benzodiazepines and related metabolites in the blood and urine. The related direct analysis in real-time tandem mass spectrometry parameters were optimized. A liquid-liquid extraction method using ethyl acetate as the extraction solvent was used for sample preparation. The established method was validated and tested on case specimens. The limits of detection of this method ranged from 0.2 to 20 ng/mL and the limits of quantification from 1 to 50 ng/mL. The recoveries ranged from 78.8% to 114%, and the matrix effects were in the range of -21.2% to 17.9%. The precision and repeatability at high and medium concentrations did not exceed 14.6%, and the limit of quantification did not exceed 18.2%, indicating a desirable linear relationship. The established method was used to determine blood and urine specimens from authentic cases, and promising results were obtained.

Possibilities and Limitations of ICP-Spectrometric Determination of the Total Content of Tin and Its Inorganic and Organic Speciations in Waters with Different Salinity Levels-Part 2: Separate Determination of Inorganic and Organic Speciations of Tin.

In this study, determination of the inorganic and organic forms of tin in waters of different salinities is considered. The possibility of the separation of speciations of tin using liquid-liquid extraction (LLE); precipitation with fluorides, iodides, ammonia, and iron (III) chloride; and sorption of organotin compounds (OTCs) was studied. The LLE and analyte precipitation methods proved to be ineffective. Inorganic and organic forms of tin were separated by the sorption of OTCs using silica gel sorbent Diapak C18. Under optimized conditions, a technique for the separate determination of the forms of tin in natural waters was developed. The technique combines hydride generation and microwave mineralization of solutions followed by ICP spectrometry. The inorganic forms of tin were determined after their solid-phase separation from organotin compounds. The lower limits of analyte quantification were 0.03 μg/L (ICP-MS) and 0.05 μg/L (ICP-OES), which provide separate determinations of inorganic and organic forms of tin in waters with different salinities. The content of OTCs in water was determined by subtracting the inorganic concentration from the total concentration of tin. The technique will allow a comprehensive assessment of the toxicological impact of tin speciations on the aquatic ecosystem.

A salting-out assisted liquid-liquid extraction method for 25 emerging pesticides in follicular fluid

Emerging pesticides of neonicotinoids (NEOs) and “Universal Pesticides” (UPs) are a growing global concern due to their growing commercial importance and potential risks to human health. The currently available analytical methods for these pesticides in biomonitoring were usually tailored for limited number of analytes, or were time consuming and costly. In this study, an efficient and sensitive method for the analysis of 16 NEOs and nine UPs in human follicular fluid (FF) was developed by using a salting-out assisted liquid-liquid extraction (SALLE) method and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Method performance was evaluated by calibration linearity (r > 0.99), sensitivity at limits of quantification (0.01–0.50 ng/mL), accuracy at relative recoveries (81–117%) and precision at relative standard deviations (≤16%). The developed method was further validated by analyzing 21 human FF samples that were collected from a hospital in Guangzhou, China. Among the 25 study analytes, two NEOs and six UPs had their detection rates over 85% and medians at 0.048–0.808 ng/mL in the FF samples. Considering the well-known toxicity of these pesticides and their metabolites, it is urgent to figure out exposure profiles of study pesticides and potential reproductive risk for women. To the best of our knowledge, this study is the first to develop and apply the SALLE method in the extraction of 16 NEOs and nine UPs simultaneously in human FF.

Assessment of Organochlorines Pesticides Residues in Water, Fish and Sediment Samples from Wurbo Lake, Bali, Taraba State, Nigeria

Organochlorines pesticides (OCPs) have been used worldwide, particularly in many African countries as in Nigeria for the control of pests. OCPs are characterized by their bioaccumulation in the environment, especially in the food chain, where they find their way into the human body. The research was aim to estimate the residual concentrations of different OCPs in water, fish and sediment from Kundi Lake to find out the extent of pesticide contamination and accumulation. The water sample was subjected to liquid-liquid extraction method while the fish and the sediment samples were subjected to soxhlet extraction. The extracts were later analysed for organochlorines pesticide residues using Gas-Chromatography coupled with mass spectrometer (GC-MS). The average length, average weight and condition factor of the studied fish species were measured. The Mormyrus rume had the highest average length of 17.12±0.05 cm, and average weight of 195.36±0.05 g follow by Clarias gariepinus which had the average length of 16.83±0.01 cm, and average weight of 189.43±0.03 g. Tilapia zilli was the least species of fish sample with a length of 12.75±0.03 cm and average weight of 102.21±0.02 g. Also, all the fish species sampled in this present study had condition factors>1. Based on the results obtained from all samples, it was found that the levels of organochlorine pesticides residues in water were generally below MRL as compared with FAO/WHO (0.5ppm). In fish samples, highest concentration of pesticide residues in Mormyrus rume follow by Clarias gariepinus, while the Tilapia zilli recorded low concentration of the pesticide residues. The Endosulfan II recorded high concentration of 4.68 ppm in Mormyrus rume follow by Endrin which recorded the concentration of 2.75 ppm, Aldrin 1.78 ppm and Atrazine 1.72 ppm all in Mormyrus rume. In Clarias gariepinus, Simazine 1.56 ppm follows by Endosulfan I 1.28 ppm while In Tilapia zilli, Endosulfan II 1.16 ppm follows by Atrazine 0.91 ppm and Beta Lindane 0.81 ppm. The results indicate that the area is contaminated with high levels of these pesticides in fish and sediment which may pose a great danger to the environment. Regular monitoring is therefore required to control the levels of pesticide residues in the water bodies.

Salting-Out-Assisted Liquid-Liquid Extraction Method for the Determination of Nicotine from Oral Traditional and Innovative Tobacco Products Using UPLC-MS/MS.

In this study, we developed and validated a novel method that allows for the extraction and quantitation of nicotine from a variety of commercially available oral tobacco products including loose and pouched traditional moist smokeless tobacco products, and oral tobacco-derived nicotine (OTDN) lozenges, gums, and pouches. The method employed an extraction technique consisting of salting-out assisted liquid-liquid extraction using sodium hydroxide and acetonitrile in conjunction with ultra-high pressure liquid chromatography coupled to mass spectrometry. Accurate quantitation was obtained using nicotine methyl-d3 isotopically labeled internal standard. Chromatographic separation of nicotine and nicotine methyl-d3 internal standard was achieved using a Waters Acquity C18 column (50 mm × 2.1 mm i.d., 2.5 μm) with 10 mM ammonium acetate buffer (pH = 10) and acetonitrile as mobile phase A and B, respectively. Using a gradient elution and a flow rate of 0.4 mL/min for 5 min runtime, nicotine eluted at 1.74 min. The method was validated according to ICH guidelines for all the sample types with an accuracy for nicotine within 89-109%. Repeatability and intermediate precision were both estimated to be ≤7% relative standard deviation (% RSD). This method is applicable for a wide range of traditional moist smokeless and OTDN tobacco products.

Investigating the effectiveness of the main channel in microfluidic liquid-liquid extraction process

The liquid-liquid extraction method has been one of the most critical chemical engineering processes for many years. The combination of liquid-liquid extraction and microfluidic systems constitutes a significant advancement in this process in recent years. The inherent advantages of microfluidic systems, due to their micro dimensions and the widespread application of liquid-liquid extraction, have made this process more applicable in various industries. This article focuses on microfluidic systems, discussing the effects of changes and modifications made to the main channel of these systems, as well as the resulting improvements in flow type and channel shape, and their applications. The assessment concludes with a discussion of the prospects of this field.