Optimization of ultrasound-assisted extraction based on response surface methodology using HPLC-DAD for the analysis of red clover (Trifolium pretense L.) isoflavones and its anti-inflammatory activities on LPS-induced 3D4/2 cell.

Introduction: Trifolium pratense L. has anti-inflammatory, antioxidant, cardiovascular disease prevention, and estrogen-like effects. The existing method for the assay of effective components is commonly based on a spectrophotometer, which could not meet the requirement of quality control. Furthermore, although there have been many studies on the anti-inflammation effect of red clover, a few have been reported on the regulatory effect of red clover isoflavones (RCI) on lipopolysaccharide (LPS)-induced inflammatory response in porcine alveolar macrophages (3D4/2 cells), and its mechanism of action is still unclear. Methods: The main components of RCI including daidzein, genistein, and biochanin A were accurately quantified by high-performance liquid chromatography coupled with diode array detection (HPLC-DAD) after optimizing the extraction process through response surface methodology. The anti-inflammatory potential of RCI was carried out by detecting the level of inflammatory cytokines and mRNA expression of related genes. Furthermore, its anti-inflammatory mechanism was explored by investigating two signaling pathways (NF-κB and MAPK). Results: The optimal extraction conditions of RCI were as follows: the concentration of ethanol is 86% and the solid-liquid ratio is 1:29, with the herb particle size of 40 mesh sieve. Under the optimal conditions, the total extraction of target components of RCI was 2,641.469 µg/g. The RCI could significantly suppress the production and expression of many pro-inflammatory cytokines. The results of the Western blot revealed that RCI dramatically reduced the expression of p65, p-p65, IκB-α, p38, and p-p38. These results are associated with the suppression of the signal pathway of p38 MAPK, and on the contrary, activating the NF-κB pathway. Collectively, our data demonstrated that RCI reversed the transcription of inflammatory factors and inhibited the expression of p65, p-p65, IκB-α, and p38, indicating that RCI had excellent anti-inflammatory properties through disturbing the activation of p38 MAPK and NF-κB pathways. Conclusion: The extraction conditions of RCI were optimized by HPLC-DAD combined with response surface methodology, which will contribute to the quality control of RCI. RCI had anti-inflammatory effects on the LPS-induced 3D4/2 cells. Its mechanism is to control the activation of NF-κB and p38 MAPK pathways, thereby reducing the expression of inflammatory-related genes and suppressing the release of cytokines.

Determination of Coenzyme Q10 Content in Food By-Products and Waste by High-Performance Liquid Chromatography Coupled with Diode Array Detection.

Coenzyme Q10 (CoQ10) is a vitamin-like compound found naturally in plant- and animal-derived materials. This study aimed to determine the level of CoQ10 in some food by-products (oil press cakes) and waste (fish meat and chicken hearts) to recover this compound for further use as a dietary supplement. The analytical method involved ultrasonic extraction using 2-propanol, followed by high-performance liquid chromatography with diode array detection (HPLC-DAD). The HPLC-DAD method was validated in terms of linearity and measuring range, limits of detection (LOD) and quantification (LOQ), trueness, and precision. As a result, the calibration curve of CoQ10 was linear over the concentration range of 1-200 µg/mL, with an LOD of 22 µg/mL and an LOQ of 0.65 µg/mL. The CoQ10 content varied from not detected in the hempseed press cake and the fish meat to 84.80 µg/g in the pumpkin press cake and 383.25 µg/g in the lyophilized chicken hearts; very good recovery rates and relative standard deviations (RSDs) were obtained for the pumpkin press cake (100.9-116.0% with RSDs between 0.05-0.2%) and the chicken hearts (99.3-106.9% CH with RSDs between 0.5-0.7%), showing the analytical method's trueness and precision and thus its accuracy. In conclusion, a simple and reliable method for determining CoQ10 levels has been developed here.

Response Surface Optimization of Dispersive Solid-Phase Extraction Combined with HPLC for the Rapid Analysis of Multiple Coccidiostats in Feed.

Since antimicrobials were banned as feed additives, coccidiostats with favorable anticoccidial action and growth promotion have been widely used in the breeding industry. The monitoring of coccidiostats in feed is necessary, while the current methods based on mass-spectrometer analysis have limited applicability and matrix effects could interfere with the results. Accordingly, in the present paper, a rapid analytical strategy for the simultaneous determination of six synthetic coccidiostats in feed using high-performance liquid chromatography coupled with diode-array detection was developed. Coccidiostats in chicken feeds were extracted with the trichloroacetic acid-acetonitrile solution. The cleanup was performed by dispersive solid-phase extraction after the optimization of the response surface methodology. The method exhibited good linearity for target coccidiostats within the range of 0.05~20 µg/mL. Recoveries for six compounds in fortified feed samples were from 67.2% to 107.2% with relative standard deviations less than 9.6%. The limit of detection was 0.2~0.3 mg/kg. The successful application of the method in commercial feed verified that it is effective and sensitive for the rapid determination of multiple coccidiostats in chicken feeds.

Analysis of active compounds and geographical origin discrimination of Atractylodes macrocephala Koidz. by using high performance liquid chromatography-diode array detection fingerprints combined with chemometrics.

In this work, a simple and effective strategy for the determination of 12 active compounds of Atractylodes macrocephala Koidz. (AM) was proposed by using high performance liquid chromatography-diode array detection (HPLC-DAD) combined with alternating trilinear decomposition (ATLD) algorithm. Utilizing the "second-order advantage", three common problems in HPLC could be resolved, namely baseline drifts, peak overlaps, and unknown interferences. 12 compounds were rapidly eluted within 12.5 min, and the average spiked recoveries were 80.8-109.9%. The figures of merit reflected the feasibility of the proposed method. Compared with the results of the traditional univariate calibration method based on HPLC-UV technique, the proposed strategy further verified the reliability and simplicity of the mathematical separation. On this basis, partial least squares-discriminant analysis (PLS-DA) was applied to discriminate 113 AM samples from different geographical origins, and variable importance in projection (VIP) was used to further screen the main differential components that affect the regional division of AM. A series of results show that the AM samples from the three regions have obviously different clustering trends. Overall, the strategy is expected to provide a scientific basis for the modern research of medicinal materials, and it is also conducive to the clinical use and market supervision of AM.

Piecewise direct standardization assisted with second-order calibration methods to solve signal instability in high-performance liquid chromatography-diode array detection systems.

In this paper, in order to solve signal instability of high-performance liquid chromatography-diode array detection (HPLC-DAD) data and maintain the second-order advantage, this work proposed piecewise direct standardization (PDS) assisted with second-order calibration methods to analyze two different complex HPLC-DAD data with signal instability, including simulated HPLC-DAD data and the data of pesticide residues in saffron. Accurate quantitative results of target analytes can be obtained by PDS combined with alternating trilinear decomposition algorithm (ATLD) and alternating trilinear decomposition assisted multivariate curve resolution algorithm (ATLD-MCR) in the presence of signal instability with time shifts and changes of peak shape. Quantitative results of the model after calibration transfer are better than those of the model established by calibration sets and prediction sets with signal instability using ATLD algorithm and ATLD-MCR algorithm under different situations. Meanwhile, t-test was used to judge whether there are significant differences between these quantitative results of models. The performances of MCR-ALS algorithm were compared with that of PDS-ATLD method and PDS-ATLD-MCR method and the proposed methods have greater potential in dealing with the case of signal instability with time shifts and changes of peak shape. In a word, this methodology can reduce the number of calibration samples for recalibration and modeling, improve the efficiency of experiment, conform to the principle of green chemistry, and obtain satisfactory quantitative results in the presence of signal instability with time shifts and changes of peak shape.

Hashish seizures in Algeria over the 2019-2020 period: The rise of potent hashish hybrids.

In Algeria, hashish is by far the most common illicit drug. This study explores Algerian hashish over a two-year period (2019-2020). A total of 2583 hashish samples from 1707 seizures for a total quantity of 108 tons were analyzed using a validated high-performance liquid chromatography-diode array detection method (HPLC-DAD). The yearly arithmetic mean tetrahydrocannabinol (THC) concentration shows relative stability-18.67% in 2019 and 19.03% in 2020 with an overall mean THC content of 18.87% and standard deviation of 10.99%. High-potency hashish (THC content > 20%) is by far the most predominant type, representing almost 50% of the total Algerian hashish seizures (mean and median around 29%). The overall mean of cannabidiol (CBD) was 2.45%, and 12% of the total studied seizures were of very low CBD concentration (CBD content<1%). Three distinct hashish chemotypes were identified: Chemotype I described the traditional Moroccan hashish with THC content ranging from 0% to 16%, Chemotype II hashish included most of the seizures and characterized by THC content ranging from 16% to 30%, and Chemotype III was characterized by hashish potency higher than 30% and very low CBD content. The identified chemotypes I, II, and III were characterized in a ternary plot, and the relative contents (THC:CBD:CBN) were about 67%:29%:4%, 88%:9%:3%, and 96%:2%:2%, respectively.

Occurrence and chemotaxonomical analysis of amatoxins in Lepiota spp. (Agaricales).

About 95% of fatal mushroom poisonings worldwide are caused by amatoxins and phallotoxins mostly produced by species of Amanita, Galerina, and Lepiota. The genus Lepiota is supposed to include a high number of species producing amatoxins. In this study, we investigated 16 species of Lepiota based on 48 recently collected specimens for the presence of amatoxins by liquid chromatography coupled to a diode-array detector and mass spectrometry (UHPLC-QTOF-MS/MS). By comparing the retention times, UV absorptions, and diagnostic MS fragment ions with data obtained from the benchmark species Amanita phalloides, we detected α-amanitin and γ-amanitin in Lepiota subincarnata, α-amanitin and amaninamide in Lepiota brunneoincarnata, and ß-amanitin and α-amanitin in Lepiota elaiophylla. Phallotoxins have not been detected any of these species. Two possibly undescribed amatoxin derivatives were found in Lepiota boudieri and L. elaiophylla, as well as one further non-amatoxin compound in one specimen of L. cf. boudieri. These compounds might be used to differentiate L. elaiophylla from L. xanthophylla and species within the L. boudieri species complex. No amatoxins were detected in L. aspera, L. castanea, L. clypeolaria, L. cristata, L. erminea, L. felina, L. fuscovinacea, L. lilacea, L. magnispora, L. oreadiformis, L. pseudolilacea, L. sp. (SeSa 5), and L. subalba. By combining the occurrence data of amatoxins with a phylogenetic analysis, a monophyletic group of amatoxin containing species of Lepiota is evident. These chemotaxonomic results highlight the relevance of systematic relationships for the occurrence of amatoxins and expand our knowledge about the toxicity of species of Lepiota.

Antiviral activity of aframomum melegueta against severe acute respiratory syndrome coronaviruses type 1 and 2.

Plant-based compounds with antiviral properties against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been identified in Aframomum melegueta through computational models. The seed extract have been traditionally used to treat different illnesses. In this study, ethanolic extracts were prepared for six commercial samples of A. melegueta seeds. Antiviral activity was tested using the XTT cytotoxicity assay and cell-based SARS-CoV-1 and 2 pseudoviral models. The presence of gingerols and other non-volatile components in the seed extracts was determined using an Agilent 1290 UPLC/DAD in tandem with an Agilent 6546 QTOF-MS. Our results showed selective antiviral activity with TI values as high as 13.1. Fifteen gingerols were identified by chromatographic analysis, with 6-gingerol being the dominant component in each seed extract. A combination of 6-gingerol with techtochrysin, previously identified in computational models as a potential active ingredient against SARS-CoV-2, demonstrated additive antiviral activity with CI values between 0.8715 and 0.9426. We confirmed the antiviral activity of A. melegueta predicted through computational models and identified a different compound, 6-gingerol, as a potential active ingredient.

Discovering the main "reinforce kidney to strengthening Yang" active components of salt Morinda officinalis based on the spectrum-effect relationship combined with chemometric methods.

Morinda officinalis, a well-known traditional herbal medicine in China, is used to treat deficiency of kidney-yang syndrome. Although this medicine has the property of "reinforcing kidney to strengthening Yang," the chemical constituents responsible for this effect remain to be elucidated. Here, we aimed to identify the main active compounds responsible for reinforcing kidney to strengthening Yang, based on spectrum-effect relationships combined with chemometrics. We used the UPLC-diode array detection method to establish the chromatography fingerprint of M. officinalis. Hydrocortisone-induced and adenine-induced kidney-yang deficiency patterns were established to evaluate the efficacy of M. officinalis. Serum triiodothyronine, free thyroxine, thyrotropin, testosterone, cortisol, luteinizing hormone, follicle-stimulating hormone, corticotropin-releasing hormone, and adrenocorticotropic hormone levels were determined as pharmacodynamic indices. Analytic hierarchy process was used to determine the weight of each index to the total pharmacodynamic contribution. Lastly, the spectrum-effect between the fingerprint and the pharmacological effects were established using grey relational analysis and partial least squares. Our findings indicated that peaks 1, 2, 3, 5, 6, 7, 8, 9, 11, 13, 15, 17, and 20 might represent the main components that positively correlated to the total effect, of which four were identified by comparison with reference standards. The identified components were monotropein (peak 1), deacetyl asperulosidic acid (peak 3), asperulosidic acid (peak 8), and asperuloside (peak 9). Our results suggest that the "reinforce kidney to strengthening Yang" effects were attributable to the combined effects of the multiple chemical components of M. officinalis and provide a valuable method to identify the active "reinforce kidney to strengthening Yang" components of M. officinalis and establish the quality control of M. officinalis.

Streamlined three step total vitamin C analysis by HILIC-UV for laboratory testing.

OBJECTIVES: In the clinical setting, the analysis and quantification of vitamin C (ascorbic acid) poses several challenges including analyte instability and poor retention by reverse phase HPLC systems. In this article we describe a rapid hydrophilic interaction chromatography ultraviolet method for the measurement of total vitamin C in plasma which overcomes these issues. METHODS: Ascorbic acid and the internal standard were separated under isocratic conditions using a Waters BEH-Amide column and a mobile phase containing 0.005 M potassium phosphate in 80% acetonitrile. RESULTS: The proposed method was validated and showed good precision (coefficient of variation <5%), accuracy (>99%), and analyte stability after extraction (>24 h). CONCLUSIONS: The simple sample preparation allows full automation and rapid analytical run times of the assay and is therefore suitable for a high-throughput clinical chromatography laboratory.

[Confirmation of unknown additives in enrofloxacin powder].

In order to ensure the safety of animal food and regulate the application of veterinary drugs, it is necessary to strictly monitor their content, and to constantly improve the methods used to detect non-specific, illegally added substances in veterinary drugs. A study about the screening, analysis, and confirmation of illegal additives in enrofloxacin powder (used for aquaculture) using non-targeted analysis technology was introduced. First, an enrofloxacin powder test solution under acidic conditions was prepared by adding formic acid, and an enrofloxacin powder test solution under alkaline conditions was prepared by adding sodium carbonate. An ultra-performance liquid chromatography with photodiode array detector (UPLC-PDA) was used to assay the test solutions for the presence of unknown additives. Results revealed two high-response unknown peaks in the acidified test solution, with retention times of 1.870 min and 5.122 min respectively. In the alkalized test solution, only one high-response unknown peak was found, with a retention time of 5.122 min. The ultraviolet spectrum characteristic peaks at 5.122 min in acidified and alkalized test solutions were similar, but the peak area in the alkalized test solution was almost ten times that in the acidified solution. Two potential unknown substances were detected. Unknown substance 1 (1.870 min) and unknown substance 2 (5.122 min) may transform under acidic or alkaline conditions. Ultra-performance liquid chromatography-time of flight high resolution mass spectrometry (UPLC-TOF-HRMS) was used to analyze the unknown compounds in more detail. The acidified and alkalized test solutions were detected in the positive and negative ion modes of mass spectrometry, respectively. Accurate mass of the precursor ion, characteristics of secondary ion fragments, and isotopic intensity ratio of the two unknown substances were collected. This information was imported into SCIEX OS software. The molecular formula of the parent ion of unknown substance 2 was found to fit to C11H8O2, and its secondary fragment structure may contain a benzene ring and two carbonyl groups, with a propylene structure connected to them through ring formation. From this, unknown substance 2 was presumed to be a menadione. The molecular ion peak of unknown substance 1 was found to fit to C11H9O5S-, only HSO3- was collected in the secondary fragments, and the missing part was consistent with unknown substance 2. Considering the most common derivatives of menadione, unknown substance 1 can be proposed to be menadione sodium bisulfite. Finally, we used menadione and menadione sodium bisulfite as reference substances in a comparative study. The same treatment method was used to prepare menadione, menadione sodium bisulfite reference solution, and enrofloxacin powder test solution. After UPLC-PDA detection, unknown substance 1 and menadione sodium bisulfite, unknown substance 2 and menadione, were found to have similar retention times and UV spectra. When the reference solution was added to the enrofloxacin powder test solution, the peak purity of the unknown substance did not change, and were all single peaks. UPLC-TOF-HRMS analysis revealed that the retention time of unknown substance 1 was consistent with that of sodium menadione bisulfite: compared to its accurate mass number in theory, the mass accuracy error was 1.0×10-6, and the matching degree of fragmentation information in the library was 100%. The retention time of unknown substance 2 was same as the menadione: compared to its accurate mass number in theory, the mass accuracy error was 0.6×10-6, and the matching degree of fragmentation information in the library was 99.7%. The structures of unknown substances 1 and 2 were confirmed. Menadione sodium bisulfite is known to participate in the synthesis of thrombin in the liver, and also promotes the formation of prothrombin, and accelerates coagulation. The indication of enrofloxacin powder (used for aquaculture) is the treatment of hemorrhage and sepsis in aquaculture animals such as fish and eel. The pharmacological effects of the two drugs correspond to each other, which can cause producers to take risks and add them illegally. With the strict supervision and severe restrictions on the addition of veterinary drugs, illegal additives are becoming more and more subtle. Conventional targeted analysis does not always meet the monitoring requirements. In this paper, the non-targeted analysis of unknown substances using UPLC-PDA combined with UPLC-TOF-HRMS is described in detail. The results may provide a technical reference for screening and identifying illegal additives in drugs, food, health care products, cosmetics, and pesticides.

[Determination of thiram in wheat flour and flour improvers by high performance liquid chromatography-diode array detection].

Thiram is an important dithiocarbamate (DTC) fungicide. In the United States and the European Union, the limit range of thiram is 0.1-15 mg/kg in fruits and vegetables, but there is no specific limit for grains. The maximum residue limit (MRL) for wheat is 1 mg/kg (calculated as carbon disulfide, CS2) in the National Food Safety Standard (GB 2763-2019). At present, the relevant regulation methods in China are targeted at the detection of dithiocarbamates and are incapable of detecting thiram specifically. CS2 is produced by the reaction of dithiocarbamate and acid, and it is then determined by spectrophotometry or GC; this renders the quantification of dithiocarbamate indirect. HPLC and HPLC-MS/MS methods are also reported for the detection of thiram. Most of the literature focuses on the determination of thiram in vegetables, fruits, soil, etc. In these methods, thiram is converted into dimethyldithiocarbamate (DMD) anions in an alkaline buffer solution, and DMD can be determined by HPLC-UV or LC-MS. However, ziram can also be converted into the DMD anion under alkaline conditions. Therefore, thiram cannot be distinguished from ziram, and this may produce false-positive results. Research has shown that in the presence of sulfite, thiram is converted into a DMD-sulfite adduct, which can be a marker for the selective determination of thiram. Furthermore, thiram can be directly detected by HPLC and HPLC-MS/MS after extraction with dichloromethane, chloroform, hexane, cyclohexane, ethyl acetate, or methanol and clean-up by solid phase extraction in vegetables and fruits. However, until now, few studies have reported the determination of thiram in wheat flour and flour improvers. Therefore, it is of great importance to develop a method for thiram in wheat flour. In this study, an analytical method based on HPLC-DAD was developed for the determination of thiram in wheat flour and flour improvers. The wheat flour and flour improver samples were extracted using acetonitrile. After shaking for 15 min, the samples were ultrasonicated for 10 min in an ice-water bath. The supernatant was filtered before separation on a ZORBAX plus-C18 column (150 mm×4.6 mm, 5 µm). The samples were eluted with a water-acetonitrile solvent system and detected at 280 nm. In this research, the extraction solvent, extraction solvent volume, ultrasonic conditions, chromatographic column, determination wavelength, and mobile phase were optimized. The retention times and UV spectra were used for qualitative analysis, and the external standard method was used to quantify thiram. Stability tests of standard stock solutions, a series of standard solutions, and extraction solutions were also performed. The standard stock solutions could be stored for at least 21 d, and the series of standard solutions could be stored for 14 d under refrigeration at 4 ℃. The standard solution was either exposed to light at room temperature for 4 h or kept in dark at room temperature for 48 h, and no obvious degradation was observed. This revealed that thiram was stable in acetonitrile solution during our investigation. It was suggested that the extraction solution should be analyzed as soon as possible. The linear range was 0.30-30.0 µg/mL. The peak area of the analyte showed a good linear relationship with its corresponding concentration, and the correlation coefficient (r2) was 0.99999. When the spiked levels were 1.5, 3.0, and 15 mg/kg, the spiked recoveries of thiram were 89.6%-98.3%, with relative standard deviations of 1.6%-3.9% (n=6). The limits of determination and quantification for thiram were 0.5 mg/kg and 1.5 mg/kg, respectively. The results revealed that this method is simple, rapid, and specific, in addition to having high precision, good repeatability, and a low limit of detection. The method is thus suitable for the daily routine analysis of thiram in wheat flour and flour improvers.

Benefits and Pitfalls of HPLC Coupled to Diode-Array, Charged Aerosol, and Coulometric Detections: Effect of Detection on Screening of Bioactive Compounds in Apples.

The new screening method for rapid evaluation of major phenolic compounds in apples has been developed. Suitability of coupling HPLC/UHPLC separation with the diode-array detection and universal charged aerosol detection with respect to the presence of interfering substances was tested. Characteristics of both detection techniques were compared and method linearity, limits of detection and quantitation, and selectivity of them determined. Student t-test based on slopes of calibration plots was applied for the detailed comparison. The diode-array detection provided the best results regarding sensitivity and selectivity of the developed method in terms of evaluation of phenolics profiles. The response of the charged aerosol detector was negatively affected by co-eluting substances during rapid-screening analyses. Coulometric detection was used for advanced characterization of extracts in terms of antioxidant content and strength to obtain more complex information concerning sample composition. This detection also allowed evaluation of unidentified compounds with antioxidant activity. HPLC/UHPLC separation using a combination of diode-array and coulometric detectors thus represented the best approach enabling quick, yet complex characterization of bioactive compounds in apples.

Effect of Crocus sativus L. Stigmas Microwave Dehydration on Picrocrocin, Safranal and Crocetin Esters.

The dehydration process is the basis to obtain high quality saffron and to preserve it for a long time. This process modifies saffron's main metabolites that define its quality, and are responsible for the characteristic color, taste, and aroma of the spice. In this work, the effect of microwave dehydration on saffron main metabolites (picrocrocin, safranal and crocetin esters) from Crocus sativus L. stigmas at three determinate powers and different time lapses was evaluated. The results showed that this dehydration process obtained similar or lower crocetin esters content, and after three months of storage, higher concentration was shown in treatments at 440 W for 36 s, 55 s, and 73 s; at 616 W for 90 s; and at 800 W for 20 s. Picrocrocin content was lower and safranal content was higher in all treatments compared to the control both before and after storage. Regarding to commercial quality, microwave dehydration obtained Category I of saffron according to International Standard Organization (ISO) 3632. After three months of storage, treatments at 616 W for 83 s and 800 W for 60 s obtained lower categories. The results obtained suggest that microwave dehydration is a suitable process for obtaining high quality saffron, 800 W with 6 lapses of 20 s being the best conditions studied.

Comparison of three chemometric methods for processing HPLC-DAD data with time shifts: Simultaneous determination of ten molecular targeted anti-tumor drugs in different biological samples.

Three intelligent chemometric multi-way calibration methods including alternating trilinear decomposition (ATLD), alternating trilinear decomposition assisted multivariate curve resolution (ATLD-MCR) and multivariate curve resolution-alternating least squares (MCR-ALS) combined with high performance liquid chromatography-diode array detection (HPLC-DAD) were used to quantify ten molecular targeted anti-tumor drugs in three complex biological matrices (plasma, urine and cell culture media matrices). All analytes can be successfully eluted in 6.5 min. In this experiment, various degrees of time shifts occurred in different samples. While slight time shifts exist in the chromatographic analysis, satisfactory results can be obtained by the three proposed methods. When the time shift was large (5.6 s), the average spiked recoveries obtained by ATLD analysis were in the range of 58.9%-116.5%, which was less than satisfactory. However, the average recoveries obtained by MCR-ALS and ATLD-MCR analysis were 89.8%-114.8% and 84.5%-106.1% respectively, and more satisfactory results were obtained. For further research, ATLD-MCR and MCR-ALS methods were compared, and the results were evaluated by statistical tests. Accuracies of concentrations obtained by them were considered to be no significant difference. In addition, compared with other methods currently published, the proposed chemometric methods combined with the HPLC-DAD can rapidly, simultaneously and accurately determine varieties of molecular targeted anti-tumor drugs in different complex biological matrices even in the presence of severe peak overlaps, severe time shifts, slight baseline drifts and different unknown background interferences.

Simultaneous analysis of thirteen phytohormones in fruits and vegetables by SPE-HPLC-DAD.

Determination of phytohormones have attracted increasing attentions in food safety field. In this study, an efficient and quantitative method was developed which can simultaneously determinate thirteen phytohormones in fruits and vegetables using solid phase extraction (SPE) combined with high performance liquid chromatography-diode array detection (HPLC-DAD). The samples were extracted with 80% methanol containing 0.5% (V/V) formic acid, and the extracts were then concentrated and purified using primary secondary amine (PSA) and C18 tandem dual SPE cartridges. The analytes were separated on a Waters XBridge™ C18 column and eluated utilizing a gradient elution program of water and methanol. Mean recoveries of the thirteen analytes varied from 74.69 to 92.40%, with relative standard deviations < 3.57%. The limits of detection and quantitation were 0.005-0.018 mg/kg and 0.02-0.10 mg/kg, respectively. The phytohormones in kiwi fruit, strawberry, bean sprout, and green pepper were detected using the above method, respectively. Only the IAA content of 0.14 mg/kg was detected for the strawberry from a supermarket, which was lower than the prescribed limit in food safety standards (0.2 mg/kg).

Quantification of letermovir in human serum using high-performance liquid chromatography with diode array detection.

A simple, rapid, cost-effective and sensitive high-performance liquid chromatography method with diode array detection was developed and validated for the quantification of letermovir, a compound approved for prophylaxis of cytomegalovirus infection and disease in adult recipients of an allogeneic hematopoietic stem cell transplant. Sorafenib was used as internal standard. Samples were pre-treated by liquid-liquid extraction with tert-butyl methylether. Separation was achieved on a XTerra® RP18 column (150 × 2.1 mm, 5 µm) at 30 °C using gradient elution with a mobile phase of 20 mM ammonium bicarbonate pH 7.9 (mobile phase A) and acetonitrile:20 mM ammonium bicarbonate (9:1 v/v) (mobile phase B). Samples were eluted at a flow rate of 0.3 mL/min throughout the 20-min run. UV wavelength mode was used, letermovir and sorafenib were monitored at 260 nm. The calibration curve was linear in a concentration range of 25-5000 ng/mL with correlation coefficients ≥ 0.99. Intra-day and inter-day accuracy expressed as relative error were -11.4-20% and -7.96-10.62%, respectively. Precision expressed as coefficient of variation was 1.44-3.15% (intra-day) and 1.17-1.93% (inter-day). The method was successfully applied for analysis of 128 letermovir levels demonstrating its usefulness for letermovir monitoring in routine clinical practice.

Effects of carrier agents on powder properties, stability of carotenoids, and encapsulation efficiency of goldenberry (Physalis peruviana L.) powder produced by co-current spray drying.

Maltodextrin, modified starch, inulin, alginate, gum arabic, and combinations thereof were used as carrier agents for spray drying of carotenoid-rich goldenberry (Physalis peruviana L.) juice and compared to cellobiose as an alternative carrier. Powders were analyzed with respect to particle size and morphology, yield, moisture content, cold water solubility, suspension stability, hygroscopicity, carotenoid encapsulation efficiency, and carotenoid retention during storage. A high initial carotenoid concentration after spray drying, a high encapsulation efficiency of 77.2%, and a slow carotenoid degradation kinetics favored the high carotenoid content of the cellobiose powder at the end of the storage. Cellobiose might protect the carotenoids from degradation processes by light exposure, high temperature, and oxygen due to a tighter particle crust and larger particle sizes. Therefore, cellobiose may be considered a potential carrier agent for the encapsulation of carotenoid-rich fruit juices.

Characterization of Algerian-Seized Hashish Over Eight Years (2011-2018). Part II: Chemical Categorization.

In Algeria, large quantities of hashish are seized every year. This study aimed to investigate the total content of major cannabinoids in the illicit seized hashish in Algeria over an 8-year period (2011-2018) in order to establish the chemical profile of North African hashish. A total of 3265 hashish samples were analyzed using a validated high-performance liquid chromatography-diode array detection (HPLC-DAD) method, allowing the simultaneous quantification of both the acidic and the neutral forms of Δ9-tetrahydrocannabinol (THC), cannabidiol (CBD), and cannabinol (CBN). The results revealed a slight upward trend in the mean THC content, from 7.0% in 2011 to 9.4% in 2018, with an overall mean value of 8.4%. The overall means of CBD and CBN content were 3.5% and 0.8%, respectively. The number of high-potency hashish samples gradually increased to reach 6% in 2018. Two distinct hashish chemotypes were identified: the highly populated chemotype II, corresponding to the traditional medium-potency hashish ([THC + CBN]/CBD ~ 2.16), and chemotype I, containing hashish samples of relatively high THC levels and low levels of CBD (ratio ~ 4.90). Both chemotypes I and II were characterized in the ternary plot, and the proportions (THC:CBD:CBN) were about 85%:13%:2% and 60%:35%:5%, respectively.

An in vitromodel to quantify interspecies differences in kinetics for intestinal microbial bioactivation and detoxification of zearalenone.

Zearalenone (ZEN) is a mycotoxin known for its estrogenic activities. The metabolism of ZEN plays a role in the interspecies differences in sensitivity to ZEN, and is known to occur in the liver and via the intestinal microbiota, although the relative contribution of these two pathways remains to be characterized. In the present study a fecal in vitro model was optimized and used to quantify the interspecies differences in kinetics of the intestinal microbial metabolism of ZEN in rat, pig and human. Vmax, Km, and catalytic efficiencies (kcat) were determined, and results obtained reveal that the kcat values for formation of α-ZEL and ß-ZEL amounted to 0.73 and 0.12 mL/h/kg bw for human microbiota, 2.6 and 1.3 mL/h/kg bw for rat microbiota and 9.4 and 6.3 mL/h/kg bw for pig microbiota showing that overall ZEN metabolism increased in the order human < rat < pig microbiota. Expressed per kg bw the kcat for ZEN metabolism by the liver surpassed that of the intestinal microbiota in all three species. In conclusion, it is estimated that the activity of the intestinal colon microbiome may be up to 36 % of the activity of the liver, and that it can additionally contribute to the species differences in bioactivation and detoxification and thus the toxicity of ZEN in pigs and rats but not in humans. The results highlight the importance of the development of human specific models for the assessment of the metabolism of ZEN.