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Summary: Neltuma ruscifolia, a neglected underutilised species (NUS) within the Fabaceae family, lacks formal commercial recognition, despite its nutritional richness and endemic status. The objective of this study was to evaluate the antioxidant and antiglycating properties, as well as to identify associated bioactive compounds, in extracts (ultrasound and agitation) of different grinding fractions of N. ruscifolia seeds. Bioactivity varied depending on the extraction procedure and milling fraction, with the highest value in ultrasound‐extracted residue (exocarp, mesocarp). Some additional polyphenolic compounds (like hydroxybenzoic/ellagic acids) only appeared upon ultrasound‐assisted extraction. It was possible to identify some polyphenols (such as chrysin, rutin, kaempferol and cinnamic, coumaric, protocatechuic, ellagic and caffeic acids) that were highly related to the bioactivity. The antioxidant, antiglycating, colourant and technological properties of the extracts different fractions of N. ruscifolia pods are mainly dependent on the selected milling fraction. The extraction method, in a second place, significantly affects the resulting quantity and the type of extracted compounds, defining their potential applications. [ABSTRACT FROM AUTHOR]

Background: Hydroxyurea (HU), also known as hydroxycarbamide, is an oral ribonucleotide reductase inhibitor. In 1999, the United States Food and Drug Administration (FDA) approved HU for the treatment of sickle cell disease (SCD). Since then, it has become the cornerstone in the management of SCD patients, helping to reduce vaso-occlusive crises, acute chest syndrome, the need for blood transfusions, hospitalizations and mortality. There is considerable variability among individuals in HU pharmacokinetic (Pk) parameters that can influence treatment efficacy and toxicity. The objective of this work is part of a clinical study aimed at investigating HU Pk and determining the optimal sampling time to estimate the Area Under the Curve (AUC) in SCD patients. Methods: HU plasma concentration in 80 patients at various time points (2, 4, 6, 24 h) following a 48-h drug washout was quantified using High-Pressure Liquid Chromatography (HPLC) coupled with an ultraviolet (UV) detection method previously described in the literature and adapted to new conditions with partial modifications. Results: The mean HU administered dose was 19.5 ± 5.1 mg/kg (range: 7.7–37.5 mg/kg). The median AUC quantified in plasma patients was 101.3 mg/L/h (Interquartile Range (IQR): 72.5–355.9) and it was not influenced by the weight-based dose. However, there was a strong positive correlation between AUC and Body Mass Index (BMI) as well as dose per Body Surface Area (BSA). Along with a three-point approach for AUC determination present in the literature, we show results obtained from a four-point sampling strategy, which is more useful and effective for better optimizing dose escalation to the maximum tolerated dose (MTD). Moreover, we observed that most patients achieved the maximum HU plasma concentration two hours after drug administration, regardless of age differences. Conclusions: HU treatment, which represents a milestone in the treatment of SCD due to its ability to reduce disease complications and improve patients' quality of life, requires careful monitoring to optimize the individual dose for saving potential side effects and/or adverse events. [ABSTRACT FROM AUTHOR]

Background: Hydroxyurea (HU), also known as hydroxycarbamide, is an oral ribonucleotide reductase inhibitor. In 1999, the United States Food and Drug Administration (FDA) approved HU for the treatment of sickle cell disease (SCD). Since then, it has become the cornerstone in the management of SCD patients, helping to reduce vaso-occlusive crises, acute chest syndrome, the need for blood transfusions, hospitalizations and mortality. There is considerable variability among individuals in HU pharmacokinetic (Pk) parameters that can influence treatment efficacy and toxicity. The objective of this work is part of a clinical study aimed at investigating HU Pk and determining the optimal sampling time to estimate the Area Under the Curve (AUC) in SCD patients. Methods: HU plasma concentration in 80 patients at various time points (2, 4, 6, 24 h) following a 48-h drug washout was quantified using High-Pressure Liquid Chromatography (HPLC) coupled with an ultraviolet (UV) detection method previously described in the literature and adapted to new conditions with partial modifications. Results: The mean HU administered dose was 19.5 ± 5.1 mg/kg (range: 7.7–37.5 mg/kg). The median AUC quantified in plasma patients was 101.3 mg/L/h (Interquartile Range (IQR): 72.5–355.9) and it was not influenced by the weight-based dose. However, there was a strong positive correlation between AUC and Body Mass Index (BMI) as well as dose per Body Surface Area (BSA). Along with a three-point approach for AUC determination present in the literature, we show results obtained from a four-point sampling strategy, which is more useful and effective for better optimizing dose escalation to the maximum tolerated dose (MTD). Moreover, we observed that most patients achieved the maximum HU plasma concentration two hours after drug administration, regardless of age differences. Conclusions: HU treatment, which represents a milestone in the treatment of SCD due to its ability to reduce disease complications and improve patients’ quality of life, requires careful monitoring to optimize the individual dose for saving potential side effects and/or adverse events.

Amikacin, an aminoglycoside, is a widely used parenteral antibiotic. Therapeutic drug monitoring (TDM) is recommended for aminoglycosides to avoid toxicity. However, the lack of infrastructure at most places precludes it. This pilot and novel study attempt to estimate the real-world serum levels of Amikacin in hospitalised patients. Thirty admitted patients, given Amikacin injections, were included in the study. In addition, 15 clinical specimens isolated with gram-negative bacteria were tested for minimum inhibitory concentration (MIC) value of Amikacin. Trough and peak serum levels of Amikacin were estimated by high-pressure liquid chromatography (HPLC). The average MIC value of Amikacin estimated in our laboratory was 3.92 mcg/mL. Peak and trough serum levels of Amikacin ranged from 12.1 to 66.4 mcg/ml and 1.1 to 20.7 mcg/ml, respectively. More than 83% of our patients achieved peak Amikacin levels of 15 mcg/mL, and 37% had trough levels above 5 mcg/mL. These levels are desirable watersheds as per available literature. Trough levels of Amikacin in all cases and a review of dosing according to MIC values are recommended to achieve drug safety and therapeutic efficacy. [ABSTRACT FROM AUTHOR]

In this work, the evaluation and comparison of mixed-mode chromatography and reversed-phase chromatography for separation of peptides and protein digests have been performed. The effects of pH of aqueous part of mobile phase as well as the effects of organic modifier on retention, resolution, and peak shape were investigated on several columns including three mixed-mode columns possessing reversed-phase/anion-exchange mechanism, two reversed-phase octadecyl columns, and one column with mixed-mode reversed-phase/anion-exchange character only in defined pH range. The set of peptides varying in their polarity, length, amino acid sequence, and charge state, namely dipeptides, N-blocked dipeptides, and oligopeptides, was selected to describe the chromatographic behavior under different conditions properly. These measurements showed the potential of mixed-mode chromatography columns for analysis of differently charged peptides in a single run. The applicability of the tested conditions has been verified by the analysis of cytochrome C digested fragments. Two types of samples were analyzed and compared, i.e., commercial cytochrome C digested standard and cytochrome C digested via trypsin spin columns. The obtained results point to the necessity of using mass spectrometry detection because of large number of unknown peaks in cytochrome C digested standard, probably originating from chymotryptic and miscleavage activities. [ABSTRACT FROM AUTHOR]

Alopecia is a pathological and multifactorial condition characterised by an altered hair growth cycle and ascribed to different pathogenic causes. Cell energetic imbalances in hair follicles occurring in this disorder could lead to the production of some "metabolic wastes", including squalene and lactic acid, which could be involved in the clinically observed sheath damage. The aim of this work was the extraction and analytical quantification of squalene and lactic acid from hair bulbs of subjects with clinical alopecia in comparison with controls, using HPLC-DAD and HPLC-MS techniques. The analytical quantification was performed after a preliminary observation through a polarised optical microscope to assess sheath damage and morphological alterations in the cases group. A significantly larger amount of squalene was quantified only in subjects affected by alopecia (n = 31) and with evident damage to hair sheaths. For lactic acid, no statistically significant differences were found between cases (n = 21) and controls (n = 21) under the experimental conditions used. Therefore, the obtained results suggest that squalene can represent a metabolic and a pathogenic marker for some alopecia conditions. [ABSTRACT FROM AUTHOR]

Understanding how natural selection acts on the genome and contributes to the process of speciation is a primary aim of the study of evolution. Here we used natural variation in two subspecies of the Guadeloupean anole (Anolis marmoratus ssp.), from the island of Guadeloupe in the Lesser Antilles, to explore the genomic basis of adaptation and speciation in Anolis lizards. These subspecies inhabit distinct ecological environments and display marked differences in adult male color and pattern. We sequenced the complete genomes of 20 anoles, 10 from each subspecies, at 1.4× coverage. We used genome‐wide scans of population differentiation, allele frequency spectrum, and linkage disequilibrium to characterize the genomic architecture within and between the subspecies. While most of the genome was undifferentiated, we observed five large divergent regions. Within these regions we identified blocks, 5 kb pairs in length, enriched for fixed single nucleotide polymorphisms. These blocks encompass 97 genes, two of which are candidate pigmentation genes. One is melanophilin (mlph), which helps transport melanosomes within melanocytes. The other is a cluster of differentiation 36 (cd36), which regulates carotenoid pigment sequestration. We used high‐pressure liquid chromatography to confirm that carotenoid pigments are significantly more abundant in the conspicuous orange‐pigmented skin of male A. m. marmoratus suggesting that cd36 may be regulating pigment deposition in this tissue. We identified for the first time a carotenoid gene that is a potential target of divergent sexual selection and may be contributing to the early stages of speciation in Anolis lizards. [ABSTRACT FROM AUTHOR]

ObjectiveWe report the effect of Hb E heterozygosity on HbA1c value by the Tosoh HLC-723G11.Case reportA 45 years-old Chinese woman presented with an abnormally low HbA1c level of 3.7% (3.9%–6.1%) in a health examination. Fasting blood glucose was normal. Blood routine examination and serum bilirubin were in the normal range. HbA1c was determined by Tosoh HLC-723G11. There was an abnormal peak between A1c and A0 on the chromatogram. Hemoglobin electrophoresis indicated that the Hb E zone accounted for 25.1%. The β-thalassemia-related genes (mutant type) were βE M/N, and the related gene CD26 (A > G) was mutated. OGTT indicated prediabetes.ConclusionHb E heterozygosity may reduce HbA1c value with abnormal chromatograms, as determined by a Tosoh HLC G11 analyzer. The Tosoh HLC G11 analyzer can well identify Hb E variation. In this case, further blood glucose-related tests should be performed to avoid missed diagnoses. However, a large sample size is needed to confirm this conclusion.

Abstract Understanding how natural selection acts on the genome and contributes to the process of speciation is a primary aim of the study of evolution. Here we used natural variation in two subspecies of the Guadeloupean anole (Anolis marmoratus ssp.), from the island of Guadeloupe in the Lesser Antilles, to explore the genomic basis of adaptation and speciation in Anolis lizards. These subspecies inhabit distinct ecological environments and display marked differences in adult male color and pattern. We sequenced the complete genomes of 20 anoles, 10 from each subspecies, at 1.4× coverage. We used genome‐wide scans of population differentiation, allele frequency spectrum, and linkage disequilibrium to characterize the genomic architecture within and between the subspecies. While most of the genome was undifferentiated, we observed five large divergent regions. Within these regions we identified blocks, 5 kb pairs in length, enriched for fixed single nucleotide polymorphisms. These blocks encompass 97 genes, two of which are candidate pigmentation genes. One is melanophilin (mlph), which helps transport melanosomes within melanocytes. The other is a cluster of differentiation 36 (cd36), which regulates carotenoid pigment sequestration. We used high‐pressure liquid chromatography to confirm that carotenoid pigments are significantly more abundant in the conspicuous orange‐pigmented skin of male A. m. marmoratus suggesting that cd36 may be regulating pigment deposition in this tissue. We identified for the first time a carotenoid gene that is a potential target of divergent sexual selection and may be contributing to the early stages of speciation in Anolis lizards.

Background: In recent years, the world has faced with the COVID-19 pandemic, followed by a significant increase in the use of antibiotics to control the COVID-19 and other secondary infections. The nonbiodegradable characteristics of antibiotics and their residues in the environment leads to increased microbial and drug resistance. Therefore, due to the high importance of antibiotics, two antibiotics, ampicillin and penicillin G, were studied in Isfahan municipal wastewater treatment plants (WWTPs). Methods: Sampling was performed for two months during 13 sampling periods and antibiotics were measured using high-performance liquid chromatography with UV detector (HPLC/UV) instrument. Results: Ampicillin and penicillin G were identified in all samples taken from the both WWTPs. The average concentration of penicillin G in WWTP E and S at the influent, effluent and its removal efficiency were 1050.54 ± 761.43 μg/L, 52.89 ± 49.27 μg/L, 89.80 ± 19.42%, 2055.12 ± 1788.08 μg/L, 143.01 ± 162.59 μg/L and 82.76 ± 21.85%, respectively. Also, the average concentration of ampicillin in WWTP E and S in the influent (796.44 ± 809.6 and 447.1 ± 322.39 μg/L), effluent (48.94 ± 24.25 and 90.31 ± 75.91 μg/L), and its removal efficiency (86.22 ± 19.84% and 66.85 ± 24.88%) were determined. Conclusion: In two studied WWTPs, the concentration of antibiotics was higher during the COVID-19 pandemic in comparison with previous studies. The statistical analysis showed that there was no significant relationship between the concentration of antibiotics in WWTPs (P < 0.05). Also, the statistical results indicated that the correlation is not significant between removal efficiency of antibiotics and removal efficiency of wastewater main parameters.

In this communication, we describe the first analytical method for the determination of free histidine in hair care products (shampoos and conditioners). Cation-exchange chromatography combined with postcolumn derivatization and fluorimetric detection enabled the accurate (recovery: 83.5–114.8%) and precise (2.4–5.6% RSD) determination of free histidine without matrix interferences at concentration levels down to 1.5 mg kg−1. Real commercially available samples were found to contain the amino acid at levels ranging between 70 and 535 mg kg−1. [ABSTRACT FROM AUTHOR]

Traditional Tibetan medicine has extensively documented the health benefits of Dracocephalum heterophyllum. However, there are few reports on the chemical composition of furanocoumarins, probably because of their complicated isolation and purification procedures. In this study, four antioxidative furanocoumarins were isolated from Dracocephalum heterophyllum by medium‐ and high‐pressure liquid chromatography in combination with on‐line high‐performance liquid chromatography−1,1‐diphenyl‐2‐picrylhydrazyl recognition. Crude samples were sequentially pretreated by medium‐pressure liquid chromatography using silica gel, MCI GEL CHP20P, and diol as stationary phases, whereas on‐line high‐performance liquid chromatography−1,1‐diphenyl‐2‐picrylhydrazyl system was used to recognize antioxidant peaks in target fractions. Thereafter, the antioxidative peaks were separated and purified through high‐pressure liquid chromatography to obtain four furanocoumarins with purities greater than 95%; namely isodemethylfuropinarine, demethylfuropinarine, alloimperatorin, and alloisoimperatorin. Finally, the antioxidant capacity of the isolated furanocoumarins was determined using in vitro experiments (1,1‐diphenyl‐2‐picrylhydrazyl assays, molecular docking, and cellular validation) and it was concluded that nuclear factor erythroid 2‐related factor 2 protein is a potential target of these compounds for their antioxidation effects. Thus, the proposed methodology exhibits excellent efficacy for the preparative isolation of high‐purity antioxidative furanocoumarins from extracts of Dracocephalum heterophyllum and it can be efficiently utilized for isolating antioxidants from other natural products. [ABSTRACT FROM AUTHOR]

Dracocephalum heterophyllum has been reported as a traditional Tibetan medicine with diverse therapeutic benefits for the effective treatment of various diseases. However, only a few reports on its free radical inhibitors are limited due to its complex chemical composition and difficult isolation and purification processes. In this study, five free radical inhibitors are isolated by an integrated chromatographic method from Dracocephalum heterophyllum, followed by an investigation of the in vitro antioxidant activity utilizing 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Medium-pressure liquid chromatography was used for the pretreatment of the crude extract of Dracocephalum heterophyllum and targeted separation and purification of the free radical inhibitors using high-pressure liquid chromatography; the antioxidant peaks are recognized by the separation and purification process in combination with an online HPLC-DPPH system. Five free radical inhibitors with purity higher than 95% were obtained, namely xanthotoxol, 5-hydroxy-8-methoxypsoralen, luteolin, methyl rosmarinate, and ethyl rosmarinate. Finally, DPPH assays are performed, and their IC50 values for isolated compounds are 250.39 ± 13.32 μM, 26.91 ± 1.93 μM, 66.87 ± 14.33 μM, 21.99 ± 3.17 μM, and 36.96 ± 1.72 μM, respectively. This method is effective in isolating free radical inhibitors from Dracocephalum heterophyllum, and it has the potential to be adopted for the isolation of antioxidants from other plants of medicinal value as well. [ABSTRACT FROM AUTHOR]

Background: Chronic kidney disease (CKD) concomitant with diabetes mellitus (DM), anemia and uremia. Thus, monitoring HbA1c levels presents a complex clinical challenge., Methods: This analytical cross-sectional study was conducted from May 2022 to April 2023 at Dr. Ram Manohar Lohia Institute of Medical Sciences, Lucknow. We compared HbA1c values obtained by the turbidimetric inhibition immunoassay (TINIA) and high-pressure liquid chromatography (HPLC) methods among non-dialysis CKD patients (n=127)., Results: HbA1c was not detectable among 27 patients by TINIA but measurable with HPLC, all being's anemic. The remaining 100 patients, it was detectable by both the methods. Among these 100 patients, linear regression analysis showed a very strong positive correlation between TINIA-HbA1c and HPLC-HbA1c (R2=0.861; p<0.0001). The agreement between methods was substantial (Cohen's kappa 0.657; p<0.0001). However, HbA1c levels were detected significantly higher with HPLC (Median 7.9, IQR 2.7) than that of TINIA (Median 7.0, IQR 2.9;p=0.025) in diabetics while the difference was not significant in non-diabetic group with both HPLC (Median 5.4,IQR 0.8) and TINIA (Median 5.1,IQR 1.1). Carbamylated Hb (CHb; as detected by HPLC as a side product) was correlated to both HbA1c by HPLC (r=0.299;p=0.007) and TINIA (r=0.336;p=0.006) as well as to serum urea levels (r=0.439;p<0.0001)., Conclusion: HPLC estimates all HbA1c patients in our study group while TINIA failed to do so in around 21.26% cases. The very low hemoglobin levels and high carbamylated hemoglobin were apparent as two most common causes. Also, the values with TINIA are significantly lower in comparison to HPLC among diabetics with CKD., Competing Interests: Declaration of Conflict of interests The authors of this article declare that there is no conflict of interest with regard to the content of this manuscript., (Copyright © 2024 International Federation of Clinical Chemistry and Laboratory Medicine (IFCC). All rights reserved.)

Alopecia is a pathological and multifactorial condition characterised by an altered hair growth cycle and ascribed to different pathogenic causes. Cell energetic imbalances in hair follicles occurring in this disorder could lead to the production of some “metabolic wastes”, including squalene and lactic acid, which could be involved in the clinically observed sheath damage. The aim of this work was the extraction and analytical quantification of squalene and lactic acid from hair bulbs of subjects with clinical alopecia in comparison with controls, using HPLC-DAD and HPLC-MS techniques. The analytical quantification was performed after a preliminary observation through a polarised optical microscope to assess sheath damage and morphological alterations in the cases group. A significantly larger amount of squalene was quantified only in subjects affected by alopecia (n = 31) and with evident damage to hair sheaths. For lactic acid, no statistically significant differences were found between cases (n = 21) and controls (n = 21) under the experimental conditions used. Therefore, the obtained results suggest that squalene can represent a metabolic and a pathogenic marker for some alopecia conditions.

Medicago sativa (alfalfa) is a widely used animal feed. However, its quality has been difficult to control due to the lack of appropriate marker compounds. Therefore, it is very necessary to select an appropriate quality marker (Q-marker) to control its quality. In this study, medium-pressure liquid chromatography and high-pressure liquid chromatography were employed to effectively prepare the separation of the Q-marker (L-tryptophan) from Medicago sativa. Firstly, using MCI GEL® CHP20P as the stationary phase, 2.5 g of the target fraction Fr3 was enriched from crude Medicago sativa extract (2.9 kg) by medium-pressure liquid chromatography. Secondly, Sephadex LH-20 was used to further separate Fr3 fractions, and the Fr34 fraction (358.3 mg) was enriched after 14 repetitions. Lastly, using the ReproSil-Pur C18 AQ preparative column, 63.4 mg of L-tryptophan was obtained by high-pressure liquid chromatography, and the purity was above 95%. The results showed that medium-pressure liquid chromatography (MCI GEL® CHP20P and Sephadex LH-20) combined with high-pressure liquid chromatography (ReproSil-Pur C18 AQ) could be used to effectively prepare the Q-marker from natural products with satisfactory purity. [ABSTRACT FROM AUTHOR]

This work deals with newly developed poly(styrene-divinylbenzene-methacrylic acid) monolithic columns for capillary electrochromatographic and capillary liquid chromatographic separations of low-molecular-weight organic compounds. The influence of the polymerization time on chromatographic properties of prepared monolithic capillary columns was studied by capillary electrochromatography and capillary liquid chromatography using a model mixture of thiourea, benzene, and its homologues. The effect of electrostatic interactions between methacrylic acid as a weak cation-exchanger and selected analytes (a mixture of three amino acids, and thiourea, or diethylamine) on their retention in capillary liquid chromatography was investigated. The determined run-to-run and batch-to-batch repeatabilities indicate high reliability of the preparation of the separation media. [ABSTRACT FROM AUTHOR]

Glycosylation is one of the most significant and abundant post-translational modifications in cells. Glycomic and glycoproteomic analyses involve the characterization of oligosaccharides (glycans) conjugated to proteins. Glycomic and glycoproteomic analysis is highly challenging because of the large diversity of structures, low abundance, site-specific heterogeneity, and poor ionization efficiency of glycans and glycopeptides in mass spectrometry (MS). MS is a key tool for characterization of glycans and glycopeptides. However, MS alone does not always provide full structural and quantitative information for many reasons, and thus MS is combined with some separation technique. This review focuses on the role of separation techniques used in glycomic and glycoproteomic analyses, liquid chromatography and capillary electrophoresis. The most important separation conditions and results are presented and discussed. [ABSTRACT FROM AUTHOR]

Today, the wide utilization of triazole fungicides due to environmental damage and its side effects has raised global concern. Thus, in this investigation, polyacrylonitrile/MnCo-layered double hydroxides nanofiber was synthesized and applied as an effective and novel adsorbent at thin-film solid-phase micro-extraction technique for the quick and concurrent extraction of five triazole fungicides in fruit and vegetable samples before quantitative analysis by high-performance liquid chromatography-ultraviolet. The incorporation of MnCo-layered double hydroxides with porous structure and abundant functional groups in a polymer medium improves the extraction efficiency of nanofibers owing to hydrogen bonding and π-π interactions formed between analytes and synthesized nano-adsorbent. Various important elements that affect the extraction efficiency of the intended analytes were optimized using a time-variable approach. Under the optimum conditions, the limit of detection and quantification range from 0.1 to 0.15 and 0.3–0.5 ng mL−1, respectively. Incorporation of the synthesized MnCo-LDH into the polymer network of PAN and the production of a novel, porous, rich in functional groups adsorbent, easy and fast synthesis by the electrospun device for the extraction of triazole fungicides in fruit and vegetable samples prior to identification by the HPLC-UV device. [Display omitted] • The MnCo-layered double hydroxides composite with a yolk-shell sphere structure. • Novel synthesis of polyacrylonitrile/MnCo-layered double hydroxides nanofibers. • Synthesized nanofibers have high absorption capacity with easy and fast synthesis. • Thin-film solid-phase micro-extraction/high-pressure liquid chromatography method. • Electrospun nanofibers with porous structure and abundant functional groups. [ABSTRACT FROM AUTHOR]

In this study, the development, validation, and application of a new liquid chromatography post-column derivatization method for the determination of Colistin in human urine samples is demonstrated. Separation of Colistin was performed using a core–shell C18 analytical column in an alkaline medium in order (i) to be compatible with the o-phthalaldehyde-based post-column derivatization reaction and (ii) to obtain better retention of the analyte. The Colistin derivative was detected spectrofluorometrically (λext/λem = 340/460 nm) after post-column derivatization with o-phthalaldehyde and N-acetyl cysteine. The post-column derivatization parameters were optimized using the Box–Behnken experimental design, and the method was validated using the total error concept. The β-expectation tolerance intervals did not exceed the acceptance criteria of ±15%, meaning that 95% of future results would be included in the defined bias limits. The limit of detection of the method was adequate corresponding to 100 nmol·L−1. The mean analytical bias (expressed as relative error) in the spiking levels was suitable, being in the range of −2.8 to +2.5% for both compounds with the percentage relative standard deviation lower than 3.4% in all cases. The proposed analytical method was satisfactorily applied to the analysis of the drug in human urine samples. [ABSTRACT FROM AUTHOR]

Childhood absence epilepsy (CAE) is characterized by absence seizures, which are episodes of lack of consciousness accompanied by electrographic spike-wave discharges. About 60% of children and adolescents with absence seizures are affected by major neuropsychological comorbidities, including anxiety. Endocannabinoids and monoamines are likely involved in the pathophysiology of these CAE psychiatric comorbidities. Here, we show that the synthetic cannabinoid receptor type 1/2 (CB1/2R) agonist WIN 55,212-2 (2 mg/kg) has a strain-dependent effect on anxiety-like and motor behavior when assess in the hole board test and cerebral monoaminergic levels in Genetic Absence Epilepsy Rats from Strasbourg (GAERS) and their non-epileptic control (NEC) rat strain. Using quantitative and Temporal pattern (T-pattern) analyses, we found that WIN 55,212-2 did not affect the emotional status of GAERS, but it was anxiolytic in NEC. Conversely, WIN 55,212-2 had a sedative effect in GAERS but was ineffective in NEC. Moreover, vehicle-treated GAERS more motivated to explore by implementing more complex and articulated strategies. These behavioral changes correlate with the reduction of 5-HT in the hippocampus and substantia nigra (SN) and noradrenaline (NA) in the entopeduncular nucleus (EPN) in vehicle-treated GAERS compared to NEC rats, which could contribute to their low anxiety status and hypermotility, respectively. On the other hand, the increased level of NA in the EPN and 5-HT in the SN is consistent with an activation of the basal ganglia output-mediated motor suppression observed in WIN 55,212-2-treated GAERS rats. These data support the view of a strain-dependent alteration of the endocannabinoid system in absence epilepsy by adding evidence of a lower emotional responsiveness and a basal ganglia hypersensitivity to cannabinoids in GAERS compared to NEC rats. [ABSTRACT FROM AUTHOR]

The health benefits of Dracocephalum heterophyllum are widely reported in traditional Tibetan medicines, but the reported chemical composition is limited, probably due to difficulties in separating and purifying compounds. In this study, antioxidative phenylethanoids were isolated from an extract of Dracocephalum heterophyllum using medium- and high-pressure liquid chromatography, coupled with on-line HPLC–1,1-diphenyl-2-picrylhydrazyl recognition. Firstly, crude samples (1.3 kg) of Dracocephalum heterophyllum were pretreated via silica gel medium-pressure liquid chromatography to yield 994.0 g of Fr2, of which 10.8 g was then pretreated via MCI GEL®CHP20P medium-pressure liquid chromatography. The resulting Fr23 and Fr25 were further separated and purified using high-pressure liquid chromatography, and yielded 8.08 mg of Fr2391, 9.76 mg of Fr2551, 16.09 mg of Fr2581, and 8.75 mg of Fr2582. Furthermore, analysis of the purity and structures of the phenylethanoids suggested that Fr2391, Fr2551, Fr2581, and Fr2582 corresponded to decaffeoylverbascoside, rosmarinic acid, acteoside, and 2′-O-acetylplantamajoside, respectively, with all being over 95% pure. Finally, the antioxidant potential of the compounds was explored based on their ability to scavenge 1,1-diphenyl-2-picrylhydrazine, as well as through molecular docking of proteins related to antioxidant pathways. Altogether, our findings revealed that the proposed method is promising for separating pure antioxidative phenylethanoids from other natural compounds. [ABSTRACT FROM AUTHOR]

Introduction: Although traditional medicines are widely used, quality and safety of some of them are not ensured due to lack of suitable quality controls and inadequate labeling. he diagnostic cellular structures and cell content for all powdered ingredients in Khurtsiin deed-6 traditional medicine were defined by "Novel" light microscopy. Biologically active compounds were identified by using TLC and HPLC. Main biologically active compounds contents were determined by UV spectrophotometer methods. Some quality and safety parameters of Khurtsiin deed-6 traditional medicine were determined by Mongolian National First Pharmacopoeia methods. Results: Some quality and safety parameters of the traditional medicine were determined as: moisture 9.63±0.09%, total ash 4.725±0.22%, and water-soluble extractive 21.28±0.11%, Alcohol soluble extractive total 46.30±0.09%, aerobic microbial count 5x10³, total yeast and mold count 3x10². The fingerprints of TLC and HPLC to reveal gallic acid, apigenin, costunolide, E-guggulsterone were defined. The total contents were measured for phenolic compounds as 65.95±1.05 mg/g and for flavonoids as 10.55 ±0.16 mg/g. Conclusion: The standardization criteria for Khurtsiin deed-6 traditional medicine were defined and Mongolian National Pharmacopeia Monograph's draft for Khurtsiin deed-6 traditional medicine was developed. [ABSTRACT FROM AUTHOR]

Childhood absence epilepsy (CAE) is characterized by absence seizures, which are episodes of lack of consciousness accompanied by electrographic spike-wave discharges. About 60% of children and adolescents with absence seizures are affected by major neuropsychological comorbidities, including anxiety. Endocannabinoids and monoamines are likely involved in the pathophysiology of these CAE psychiatric comorbidities. Here, we show that the synthetic cannabinoid receptor type 1/2 (CB1/2R) agonist WIN 55,212-2 (2 mg/kg) has a strain-dependent effect on anxiety-like and motor behavior when assess in the hole board test and cerebral monoaminergic levels in Genetic Absence Epilepsy Rats from Strasbourg (GAERS) and their non-epileptic control (NEC) rat strain. Using quantitative and Temporal pattern (T-pattern) analyses, we found that WIN 55,212-2 did not affect the emotional status of GAERS, but it was anxiolytic in NEC. Conversely, WIN 55,212-2 had a sedative effect in GAERS but was ineffective in NEC. Moreover, vehicle-treated GAERS more motivated to explore by implementing more complex and articulated strategies. These behavioral changes correlate with the reduction of 5-HT in the hippocampus and substantia nigra (SN) and noradrenaline (NA) in the entopeduncular nucleus (EPN) in vehicle-treated GAERS compared to NEC rats, which could contribute to their low anxiety status and hypermotility, respectively. On the other hand, the increased level of NA in the EPN and 5-HT in the SN is consistent with an activation of the basal ganglia output-mediated motor suppression observed in WIN 55,212-2-treated GAERS rats. These data support the view of a strain-dependent alteration of the endocannabinoid system in absence epilepsy by adding evidence of a lower emotional responsiveness and a basal ganglia hypersensitivity to cannabinoids in GAERS compared to NEC rats.

Background: The present work is focused on developing a simple stability indicating method for the estimation of the assay of benzyl alcohol and glycopyrrolate in glycopyrrolate injection using reverse high performance liquid chromatography. Methods: A simple, selective, rapid, precise, and gradient reversed-phase high-pressure liquid chromatography procedure has been proposed to estimate the assay of benzyl alcohol and glycopyrrolate for glycopyrrolate injection. It was developed on a Kromasil 100-5, C8 250 mm x 4.6 mm, 5 µm column using mobile phase-A, containing pH 2.5 potassium dihydrogen phosphate buffer and methanol in a ratio of 80:20 v/v. and mobile phase-B, containing pH 2.5 potassium dihydrogen phosphate buffer and methanol in a ratio of 10:90 v/v at a flow rate of 1.5 mL/min. The detection of glycopyrrolate and benzyl alcohol was carried out at 222 nm and 254 nm. A sharp peak was obtained for glycopyrrolate and benzyl alcohol at a retention time of about 8.4 and 4.8 min. Results: The method was validated for specificity (blank, system suitability solution, standard, and sample and stress samples for any interference), precision (six replicate sample preparations), robustness (by slightly varying critical method parameters), and solution stability, and the results were found satisfactory. Conclusion: The method is specific, precise, accurate, and robust for estimation of the assay of benzyl alcohol of glycopyrrolate in glycopyrrolate injection. [ABSTRACT FROM AUTHOR]

Abstract Background The aim of the present work was to determine potential toxicity of degradation products of febantel generated under different stress conditions mentioned in guideline Q1A (R2) laid down by International Council for Harmonization (ICH). The stability behavior of febantel was studied by subjecting it to hydrolytic, oxidative, photolytic and thermal forced degradation conditions. Results Five degradation products (DPs) were observed which were resolved using high-performance liquid chromatography (HPLC) and characterized by LC-MS/MS using positive mode of electrospray ionization. The chromatographic separation was carried out on Hypersil® BDS C18 (150 × 4.6 mm, 5 μm) column. Optimum resolution was obtained using ammonium formate buffer (10 mM, pH 3.5) and acetonitrile programmed in gradient elution mode at 281.0 nm using photodiode array detector. Conclusion The drug was found susceptible to degradation under all the stress conditions except thermal and oxidative stress. Five major unknown degradation products DP–I, DP–II, DP–III, DP–IV, and DP–V generated under photolytic, alkali, and acidic stress condition were identified and characterized by LC-MS/MS. The drug and identified degradation products were screened for prediction of in-silico toxicity using software viz. Swiss ADME, OSIRIS Property Explorer and Pro Tox II which indicated overall no toxicological concerns. Graphical abstract

Background: Propylthiohinothiadiazole is a thiadiazole derivative with pronounced antimycobacterial action. Substance is researched in order to develop methods of analysis and standardization, analytical documents regulating quality for use as an active principle in the development of pharmaceutical forms. Material and methods: 3 series of propylthiohinothiadiazole, internal reference standard of 2-(propylthio)-5H-[1,3,4]-thiadiazole[2,3-b]-quinazolin5-one substance with concentration 99.98%, chromatographic system Shimadzu LC-20AD high-pressure liquid chromatography (HPLC) with UV-VIS detector, acetonitrile and dimethylsulfoxide of purity grade “pro HPLC analysis” (Sigma Aldrih and Merck), bidistilled purified water. Results: Linearity is proved for concentrations of 5-30 µg/ml, the linear regression equation is y=36472x-44580, R²=0.9999. The limit of detection is 0.729 µg/ml and the limit of quantification is 2.210 µg/ml. It was established that the method is accurate (mean recovery values at 80%, 100% and 120% concentration levels were close to 100%). The accuracy of the method was expressed by repeatability and intermediate accuracy. The variation of the chromatographic conditions established that the method is robust. For all validation parameters, relative standard deviation was less than 1. Conclusions: The validation results show that the developed HPLC method is simple, fast, accurate and reproducible.

Abstract Background Low concentrations of morphine are required for safe dosing for intrathecal injections. Sometimes, manual dilution of morphine is performed to achieve these low concentrations, but risks dilution errors and bacterial contamination. The primary goal was to compare the concentrations of morphine and bupivacaine between four groups of syringes. The secondary goal was to investigate the difference in contamination rate between these groups. Methods Twenty-five experienced anesthesia providers were asked to prepare a mixture of bupivacaine 2.0 mg/ml and morphine 60 μg/ml using 3 different methods as clean and precise as possible. The fourth method used was the aspiration of ampoules prepared by the pharmacy. The concentrations of morphine and bupivacaine were measured by High-Pressure Liquid Chromatography (HPLC). The medication was cultured for bacterial contamination. Results Group 1 (median 60 μg/ml; 95% CI: 59–110 μg/ml) yielded 3 outliers above 180 μg/ml morphine concentration. Group 2 (76 μg/ml; 95% CI: 72–80 μg/ml) and 3 (69 μg/ml; 95% CI: 66–71 μg/ml) were consistently higher than the target concentration of 60 μg. The group “pharmacy” was precise and accurate (59 μg/ml; 95% CI: 59–59 μg/ml). Group 2 and “pharmacy” had one contaminated sample with a spore-forming aerobic gram-positive rod. Conclusion Manually diluted morphine is at risk for deviating concentrations, which could lead to increased side-effects. Medication produced by the hospital pharmacy was highly accurate. Furthermore, even when precautions are undertaken, contamination of the medication is a serious risk and appeared to be unrelated to the dilution process.

In this communication, we describe the first analytical method for the determination of free histidine in hair care products (shampoos and conditioners). Cation-exchange chromatography combined with postcolumn derivatization and fluorimetric detection enabled the accurate (recovery: 83.5–114.8%) and precise (2.4–5.6% RSD) determination of free histidine without matrix interferences at concentration levels down to 1.5 mg kg−1. Real commercially available samples were found to contain the amino acid at levels ranging between 70 and 535 mg kg−1.

Background: In a low and middle-income country (LMIC) like India, non—communicable diseases (NCDs) contribute a major proportion (61.8%) of all causes of death. Out of this, 48% of cardiovascular diseases, 23% of Chronic Respiratory Diseases, and 10% of Cancer deaths are attributable to tobacco use. Tobacco use is a major risk factor for NCDs and thus, the tobacco cessation approach is a high priority intervention to combat complications and death among NCD patients. While several interventions are available for tobacco cessation, in resource constraint countries like India, the effectiveness of low-cost, culturally specific patient-centric tobacco cessation behavioral intervention holds a potential that needs to be evaluated. A newly developed evidence-based tobacco cessation intervention package will be compared with the existing/usual care provided under the National Programme for Prevention and Control of Cancers, Diabetes, Cardiovascular Diseases, and Stroke (NPCDCS) at NCD clinics. Methods and design: 2 arm, parallel-group randomized controlled trial. Participants: Patients aged ≥30 years suffering from NCD, currently using tobacco, and attending NCD clinics in 2 districts of Punjab, India. Sample size: A total of 200 participants meeting the selection criteria will be recruited. They will be allocated either to the intervention arm or control (usual care) arm (100 each) using block randomization. Intervention: For the participants, there will be 4 face-to-face disease-specific cessation counseling sessions, disease-specific pamphlets, short text messages in vernacular language, that is, Punjabi. Follow-ups will be done at the third, sixth, ninth, and 12th months. Primary outcome: Seven-day abstinence, biochemically verified by plasma cotinine levels. Secondary outcome: Quit attempts, number of sticks/number of times of smokeless tobacco usage in a day, and stage of behavior change in tobacco users. Discussion: This multicomponent culturally specific-patient-centric behavioral intervention package for tobacco cessation at NCD clinic settings focusing on the individual, family, and social environment could increase the outreach of cessation services using existing resources, thereby strengthening health systems and enhancing the quality of life of NCD patients. Trial registration: The protocol for the study has been registered with the Clinical Trials Registry in India under the registration number CTRI/2018/01/011643. [ABSTRACT FROM AUTHOR]

Targeted cancer treatment is a promising, less invasive alternative to chemotherapy as it is precisely directed against tumor cells whilst leaving healthy tissue unaffected. The plant-derived enzyme horseradish peroxidase (HRP) can be used for enzyme prodrug cancer therapy with indole-3-acetic acid or the analgesic paracetamol (acetaminophen). Oxidation of paracetamol by HRP in the presence of hydrogen peroxide leads to N-acetyl-p-benzoquinone imine and polymer formation via a radical reaction mechanism. N-acetyl-p-benzoquinone imine binds to DNA and proteins, resulting in severe cytotoxicity. However, plant HRP is not suitable for this application since the foreign glycosylation pattern is recognized by the human immune system, causing rapid clearance from the body. Furthermore, plant-derived HRP is a mixture of isoenzymes with a heterogeneous composition. Here, we investigated the reaction of paracetamol with defined recombinant HRP variants produced in E. coli, as well as plant HRP, and found that they are equally effective in paracetamol oxidation at a concentration ≥ 400 µM. At low paracetamol concentrations, however, recombinant HRP seems to be more efficient in paracetamol oxidation. Yet upon treatment of HCT-116 colon carcinoma and FaDu squamous carcinoma cells with HRP–paracetamol no cytotoxic effect was observed, neither in the presence nor absence of hydrogen peroxide. [ABSTRACT FROM AUTHOR]

Dracocephalum heterophyllum has been reported as a traditional Tibetan medicine with diverse therapeutic benefits for the effective treatment of various diseases. However, only a few reports on its free radical inhibitors are limited due to its complex chemical composition and difficult isolation and purification processes. In this study, five free radical inhibitors are isolated by an integrated chromatographic method from Dracocephalum heterophyllum, followed by an investigation of the in vitro antioxidant activity utilizing 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay. Medium-pressure liquid chromatography was used for the pretreatment of the crude extract of Dracocephalum heterophyllum and targeted separation and purification of the free radical inhibitors using high-pressure liquid chromatography; the antioxidant peaks are recognized by the separation and purification process in combination with an online HPLC-DPPH system. Five free radical inhibitors with purity higher than 95% were obtained, namely xanthotoxol, 5-hydroxy-8-methoxypsoralen, luteolin, methyl rosmarinate, and ethyl rosmarinate. Finally, DPPH assays are performed, and their IC50 values for isolated compounds are 250.39 ± 13.32 μM, 26.91 ± 1.93 μM, 66.87 ± 14.33 μM, 21.99 ± 3.17 μM, and 36.96 ± 1.72 μM, respectively. This method is effective in isolating free radical inhibitors from Dracocephalum heterophyllum, and it has the potential to be adopted for the isolation of antioxidants from other plants of medicinal value as well.

In this work, effects of particle and ligand types on retention and separation of sets of neutral (naphthoflavones), acidic (profens), basic (cathinones), and zwitterionic (dipeptides) biologically active compounds are shown in detail and discussed. Nine columns differing in ligand and particle types were tested, namely XSelect HSS C18, XSelect HSS C18 SB, XSelect HSS PFP, XSelect CSH C18, XSelect CSH Phenyl-Hexyl, XSelect CSH Fluoro-Phenyl, XBridge C18, XBridge Shield RP18, and Atlantis PREMIER BEH C18 AX. Various mobile phases differing in acetonitrile/aqueous part volume ratio and aqueous part pH, i.e. pH = 2.1, pH = 4.7, and pH = 8.0 were tested and compared in terms of analytes´ retention and separation. Results showed significant effect of ionic interactions resulting from particle types used or additional ionisable functional groups bonded to the surface on retention and separation of charged compounds. All tested sets of compounds were baseline separated but different conditions, that is column type and mobile phase composition, were needed. [ABSTRACT FROM AUTHOR]

• Evaluation of hemoglobin variants using capillary electrophoresis by Sebia provides comparable information to high-pressure liquid chromatography by Primus. • Reference intervals for Hb A, A 2 and F in normal individuals agreed between the methods. • Common hemoglobin variants containing Hb S and Hb C were detected by both methods. • The use of a secondary method employing a different technique improves the likelihood of correct variant identification. Hemoglobin fractionation by capillary zone electrophoresis (CE) is becoming a popular method for the identification of hemoglobin variants that can cause hemoglobinopathies. The goal of this study was to compare the performance of capillary electrophoresis using Sebia Capillarys 2 Flex Piercing system (CE-S) with high-pressure liquid chromatography (HPLC) using Primus Ultra2 Resolution Variants System (HPLC-P) as a primary method in hemoglobinopathy work-up. A total of 306 blood specimens submitted for evaluation of hemoglobinopathies were studied using HPLC-P and CE-S. The reference ranges for Hb A, A 2 and F agreed well between methods. All common variants containing Hb S and Hb C were detected by both methods. Quantification of Hb A 2 with HPLC-P required a correction in the presence of Hb S, while quantification of Hb A 2 was slightly overestimated by CE-S in the presence of Hb C. Of 41 samples containing other variants, 2 were not identified by HPLC-P and 3 were not identified by CE-S. CE-S provides comparable information to that obtained by HPLC-P and it is a reliable primary method for the evaluation of hemoglobin variants. [ABSTRACT FROM AUTHOR]

Aim: This pilot cross-sectional study focused on biological monitoring of the benzene, toluene, ethylbenzene, and xylene (BTEX) urinary metabolites trans, trans-muconic acid (tt-MA), s-phenyl mercapturic acid (SPMA), hippuric acid (HA), mandelic acid (MA), and methylhippuric acid (MHA) and measured the effects of workplace BTEX exposure on pulmonary function of workers engaged in footwear manufacturing. Materials and Methods: Urinary metabolites tt-MA, SPMA, HA, MA, and MHA concentration in urine samples of study participants (N = 35) were analyzed by reverse-phase high-pressure liquid chromatography. Pulmonary function parameters were measured by spirometer and peak flow meter. Demographic information and work exposure information of study participants were collected by questionnaire interview. Results: In exposed workers, concentration of urinary SPMA, tt-MA, and HA was significantly higher (P < 0.01, in all) than the control group. Reduction in both force expiratory volume in 1 s (Forced expiratory volume in one second) and peak expiratory flow rate (PEFR) were inversely associated (P < 0.01) with growing years of age among all workers. Inverse association was measured between urinary tt-MA and decline in forced vital capacity and PEFR (P < 0.05, for each) in workers. Based on the questionnaire interview, workers were not found to be aware of workplace exposure hazards. Conclusions: Exposure biomarkers of benzene (tt-MA and SPMA) and toluene (HA) were significantly higher in workers than the control group. Study results evident the presence of occupational exposure to benzene and toluene in footwear workers. Deterioration in FEV1 and PEFR were also measured among all workers with growing years of age. The sample size was small in the present study, so further research required to confirm our results. [ABSTRACT FROM AUTHOR]

Medicago sativa (alfalfa) is a widely used animal feed. However, its quality has been difficult to control due to the lack of appropriate marker compounds. Therefore, it is very necessary to select an appropriate quality marker (Q-marker) to control its quality. In this study, medium-pressure liquid chromatography and high-pressure liquid chromatography were employed to effectively prepare the separation of the Q-marker (L-tryptophan) from Medicago sativa. Firstly, using MCI GEL® CHP20P as the stationary phase, 2.5 g of the target fraction Fr3 was enriched from crude Medicago sativa extract (2.9 kg) by medium-pressure liquid chromatography. Secondly, Sephadex LH-20 was used to further separate Fr3 fractions, and the Fr34 fraction (358.3 mg) was enriched after 14 repetitions. Lastly, using the ReproSil-Pur C18 AQ preparative column, 63.4 mg of L-tryptophan was obtained by high-pressure liquid chromatography, and the purity was above 95%. The results showed that medium-pressure liquid chromatography (MCI GEL® CHP20P and Sephadex LH-20) combined with high-pressure liquid chromatography (ReproSil-Pur C18 AQ) could be used to effectively prepare the Q-marker from natural products with satisfactory purity.

The determination of the monomer fractions in polyhydroxyalkanoates is of great importance for research on microbial-produced plastic material. The development of new process designs, the validation of mathematical models, and intelligent control strategies for production depend enormously on the correctness of the analyzed monomer fractions. Most of the available detection methods focus on the determination of the monomer fractions of the homopolymer poly(3-hydroxybutyrate). Only a few can analyze the monomer content in copolymers such as poly(3-hydroxybutyrate-co-3-hydroxyvalerate), which usually require expensive measuring devices, a high preparation time or the use of environmentally harmful halogenated solvents such as chloroform or dichloromethane. This work presents a fast, simple, and inexpensive method for the analysis of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with high-performance liquid chromatography. Samples from a bioreactor experiment for the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) with Cupriavidus necator H16 were examined regarding their monomer content using the new method and gas chromatography analysis, one of the most frequently used methods in literature. The results from our new method were validated using gas chromatography measurements and show excellent agreement. Key points ∙ The presented HPLC method is an inexpensive, fast and environmentally friendly alternative to existing methods for quantification of monomeric composition of PHBV. ∙ Validation with state of the art GC measurement exhibits excellent agreement over a broad range of PHBV monomer fractions. [ABSTRACT FROM AUTHOR]

In this study, the development, validation, and application of a new liquid chromatography post-column derivatization method for the determination of Colistin in human urine samples is demonstrated. Separation of Colistin was performed using a core–shell C18 analytical column in an alkaline medium in order (i) to be compatible with the o-phthalaldehyde-based post-column derivatization reaction and (ii) to obtain better retention of the analyte. The Colistin derivative was detected spectrofluorometrically (λext/λem = 340/460 nm) after post-column derivatization with o-phthalaldehyde and N-acetyl cysteine. The post-column derivatization parameters were optimized using the Box–Behnken experimental design, and the method was validated using the total error concept. The β-expectation tolerance intervals did not exceed the acceptance criteria of ±15%, meaning that 95% of future results would be included in the defined bias limits. The limit of detection of the method was adequate corresponding to 100 nmol·L−1. The mean analytical bias (expressed as relative error) in the spiking levels was suitable, being in the range of −2.8 to +2.5% for both compounds with the percentage relative standard deviation lower than 3.4% in all cases. The proposed analytical method was satisfactorily applied to the analysis of the drug in human urine samples.

The health benefits of Dracocephalum heterophyllum are widely reported in traditional Tibetan medicines, but the reported chemical composition is limited, probably due to difficulties in separating and purifying compounds. In this study, antioxidative phenylethanoids were isolated from an extract of Dracocephalum heterophyllum using medium- and high-pressure liquid chromatography, coupled with on-line HPLC–1,1-diphenyl-2-picrylhydrazyl recognition. Firstly, crude samples (1.3 kg) of Dracocephalum heterophyllum were pretreated via silica gel medium-pressure liquid chromatography to yield 994.0 g of Fr2, of which 10.8 g was then pretreated via MCI GEL®CHP20P medium-pressure liquid chromatography. The resulting Fr23 and Fr25 were further separated and purified using high-pressure liquid chromatography, and yielded 8.08 mg of Fr2391, 9.76 mg of Fr2551, 16.09 mg of Fr2581, and 8.75 mg of Fr2582. Furthermore, analysis of the purity and structures of the phenylethanoids suggested that Fr2391, Fr2551, Fr2581, and Fr2582 corresponded to decaffeoylverbascoside, rosmarinic acid, acteoside, and 2′-O-acetylplantamajoside, respectively, with all being over 95% pure. Finally, the antioxidant potential of the compounds was explored based on their ability to scavenge 1,1-diphenyl-2-picrylhydrazine, as well as through molecular docking of proteins related to antioxidant pathways. Altogether, our findings revealed that the proposed method is promising for separating pure antioxidative phenylethanoids from other natural compounds.

Lorcaserin is a preferential serotonin2C receptor (5-HT2CR) agonist effective to treat obesity that has also recently been proposed to treat addiction and epilepsy. Central dopamine (DA) mechanisms are likely involved in the lorcaserin mechanism of action, but other monoamines 5-HT and noradrenaline (NA) contents or their interaction with DA might account for its effects. Here we showed that lorcaserin at 3, but not 0.3 mg/kg enhanced 5-HT content in the insular cortex, the core of the nucleus accumbens, and ventral hypothalamus. Without affecting the metabolite 5-hydroxy indole acetic acid, lorcaserin reduced the indirect index of 5-HT turnover in the hippocampus, substantia nigra, and habenula. Lorcaserin at 3 mg/kg increased NA content in the orbitofrontal cortex, the central amygdala (also at 0.3 mg/kg), the ventral hypothalamus, and the shell of the nucleus accumbens. A correlative analysis of the tissue contents between pairs of brain regions revealed that 0.3 mg/kg lorcaserin enhanced the number of correlations for 5-HT, its metabolism, and NA to a lower extent. The correlation profiles were very different between saline, 0.3 and 3 mg/kg lorcaserin. Lorcaserin enhanced the correlations established between NA or 5-HT at 0.3 and 3 mg/kg and reduced the number of correlations established between the index of the turnover for DA and 5-HT. These results show that lorcaserin modulates the biochemistry of NA and 5-HT systems in a subset of brain regions. Qualitatively, they reveal, oppositely to the DA changes, that lorcaserin at 0.3, but not 3 mg/kg, enhanced the number of correlations of 5-HT content between brain regions.

Oligomeric proanthocyanidins are among the most widely distributed polyphenols in the plant kingdom, being endowed with antioxidant, anti-inflammatory, antimicrobial, immunomodulatory, antitumor and cardioprotective activities. Since oligomeric proanthocyanidins are found in limited amounts in plants and plant-derived products, there is a great need for efficient separation methodologies that can supply analytical quality standards or sufficient material for further bioassays. By efficiently providing a good molar mass distribution of oligomeric proanthocyanidins according to their degree of polymerization, Sephadex LH is the most used sorbent in column chromatography, together with other size exclusion polymers, such as Toyopearl, Diaion, MCI gel CHP or Amberlite. Combination of normal and reversed-phase high pressure liquid chromatography can significantly reduce the total number of chromatographic steps required to obtain high purity oligomeric proanthocyanidins, whereas hydrophilic interaction liquid chromatography on Diol phases can yield proanthocyanidins with a higher degree of polymerization. By lacking a solid support, countercurrent separations can solve issues related to irreversible adsorption, giving higher recoveries, low sample denaturation and a facile scale-up from analytical to industrial production. By taking advantage of the complementarity of all these chromatographic techniques, high-purity oligomeric proanthocyanidin fractions or individual compounds can be obtained on a large scale in a reduced number of separation steps. [ABSTRACT FROM AUTHOR]

The powder bed-based 3D printing process called multi-jet fusion from HP is used in various industries for different applications. A polymer powder, the so-called fusing and detailing agents, and an IR source are needed. These novel materials offer some challenges for analytical characterization. High-resolution mass spectrometry in combination with a data base search was used as a tool for comprehensive characterization of the agents, the methanol extract of new polyamide 12 powder, and the methanol extract of a grinded 3D printed part. In addition, thermogravimetric studies were carried out. Besides biocides and substances containing ethoxy units, 2-pyrrolidone and triethylene glycol were identified as organic solvents in the agents. The detailing agent contained 3.7% of 2-pyrrolidone and 11.1% of triethylene glycol. The fusing agent contained 18.7% of 2-pyrrolidone and 8.4% of triethylene glycol. Based on the agents' different functionalities, the water content of the detailing agent was 18% higher as compared to the fusing agent. In addition, the fusing agent contained a carbon black amount of 5.2%. The methanol extract of new polyamide 12 powder contained remaining monomer and oligomer compounds, as well as the oxidation stabilizer Irganox 1098. In the methanol extract of the grinded 3D printed part the cyclic and linear monomer, oligomers, and oxidation stabilizer were found. Moreover, ingredients of the agents were detected. Residues of 2-pyrrolidone, triethylene glycol, 3-benzisothiazolinone, and substances containing ethoxy units were identified in the methanol extract of the grinded 3D printed part. In contrast, no 2-methyl-4-isothiazolin-3-one was observed. [ABSTRACT FROM AUTHOR]

Background: The purpose of this study was to investigate the bioavailability enhancing potential of resveratrol (RSV) on diltiazem (DLT) treatment in healthy human volunteers. Materials and Methods: An open-label, two-period, sequential study was conducted in 12 healthy human male volunteers. A single dose of RSV 500 mg was administered daily for 10 days during the treatment phase. A single dose of DLT 30 mg was administered during the control phase and after treatment phases under fasting conditions. The blood samples were collected at predetermined time intervals after DLT dosing and analyzed using high-performance liquid chromatography. Results: Treatment with RSV significantly enhanced maximum plasma concentration, area under the curve, whereas the volume of distribution and apparent clearance (CL/F) of DLT was significantly decreased when compared to control. Conclusions: The results suggested that enhanced bioavailability of DLT might be attributed by bio enhancing the potential of RSV resulted by inhibition of P-gp and CYP3A4. Further, the dosage of DLT should be readjusted when it is used concomitantly with RSV supplements or food containing RSV. [ABSTRACT FROM AUTHOR]

Lorcaserin is a preferential serotonin2C receptor (5-HT2CR) agonist effective to treat obesity that has also recently been proposed to treat addiction and epilepsy. Central dopamine (DA) mechanisms are likely involved in the lorcaserin mechanism of action, but other monoamines 5-HT and noradrenaline (NA) contents or their interaction with DA might account for its effects. Here we showed that lorcaserin at 3, but not 0.3 mg/kg enhanced 5-HT content in the insular cortex, the core of the nucleus accumbens, and ventral hypothalamus. Without affecting the metabolite 5-hydroxy indole acetic acid, lorcaserin reduced the indirect index of 5-HT turnover in the hippocampus, substantia nigra, and habenula. Lorcaserin at 3 mg/kg increased NA content in the orbitofrontal cortex, the central amygdala (also at 0.3 mg/kg), the ventral hypothalamus, and the shell of the nucleus accumbens. A correlative analysis of the tissue contents between pairs of brain regions revealed that 0.3 mg/kg lorcaserin enhanced the number of correlations for 5-HT, its metabolism, and NA to a lower extent. The correlation profiles were very different between saline, 0.3 and 3 mg/kg lorcaserin. Lorcaserin enhanced the correlations established between NA or 5-HT at 0.3 and 3 mg/kg and reduced the number of correlations established between the index of the turnover for DA and 5-HT. These results show that lorcaserin modulates the biochemistry of NA and 5-HT systems in a subset of brain regions. Qualitatively, they reveal, oppositely to the DA changes, that lorcaserin at 0.3, but not 3 mg/kg, enhanced the number of correlations of 5-HT content between brain regions. [ABSTRACT FROM AUTHOR]

Objective: A novel stability-indicating high-pressure liquid chromatography procedure was developed followed by validation for the simultaneous estimation of allopurinol and lesinurad in bulk and was applied on the marketed formulations of allopurinol and lesinurad. Materials and Methods: The mobile phase used for the chromatographic runs consisted of methanol: acetonitrile:0.05 M phosphate buffer, pH 6.2 (25:60:15 v/v/v). The separation was achieved on a ProntoSIL Hypersorb ODS C18 column (4.6 mm x250 mm; 5 µ) using isocratic mode. Allopurinol and lesinurad peaks are well separated and detected at 239 nm using ultraviolet detector. Results: The method was linear in the quantity range of 75-600 µg/ml for allopurinol and 50-400 µg/ml for lesinurad. The method was verified in full compliance with ICH directives. Allopurinol's detection limit (DL) and quantification limit (QL) were 0.032 µg/ml and 0.097 µg/ml, respectively, while DL and QL for lesinurad were 0.022 µg/ml and 0.067 µg/ml, respectively. The validation study results showed that developed procedure was selective (no obstruction from diluent and excipients of tablet), specific (no obstruction from degradants formed during stability testing), precise (relative standard deviation [RSD] value - <2.0%), rugged (RSD value - <2.0%), accurate (recovery percent near to 100%), and robust (difference value - <2.0%). Conclusion: The reported method can also be used effectively in marketed formulations for routine analysis of allopurinol and lesinurad. [ABSTRACT FROM AUTHOR]

R-phycoerythrin is a fluorescent red protein-pigment complex present in red algae. It belongs to the light-harvesting phycobiliprotein family with phycocyanin and allophycocyanin, which together make up the phycobilisome. The protein is most commonly used for its fluorescence properties in biochemistry. To determine the highest R-phycoerythrin yielding red algal species, a reliable quantification method is required. Calculating R-phycoerythrin yield from the crude extract's absorption values has been the quickest and most widely used technique but at the same time, often misleading approach when impurities are present. Small phycoerythrin, which is often present in red algal suspensions, alters the absorption curve and leads to erroneous results. In the given study, we utilized an HPLC-SEC method with fluorescence and photo-diode array detectors, which is capable of separating the phycobiliproteins from interfering compounds. Also, the technique can detect and quantitate R-phycoerythrin yields reliably. A total of nine red algal species from Estonian and Japanese waters were tested, and small phycoerythrin was detected from most of the samples. For two of the tested species, the spectrophotometric quantification methods overestimated the yields by more than 200%. [ABSTRACT FROM AUTHOR]

Abstract: The objective of this work was to validate a fast method with low-solvent use, for the analysis of trans-resveratrol and quercetin in red wines. The wines were prepared, using a classical method, from the grape (Vitis vinifera) cultivars Cabernet Franc, Cabernet Sauvignon, Malbec, Merlot, Petit Verdot, Pinot Noir, Ruby Cabernet, Syrah, Tannat, and Tempranillo. Samples were filtered and analyzed by high-pressure liquid chromatography (HPLC) with a diode array detector (DAD), at 306 and 371 nm, for trans-resveratrol and quercetin, respectively. An octylsilane column was used, and the mobile phase was composed by a gradient of methanol, water, and formic acid. The method was validated according to the following figures of merit: specificity, linearity, limit of detection, limit of quantification, precision, accuracy, and robustness. The proposed HPLC-DAD method may be established for the analysis of trans-resveratrol and quercetin in red wines.