Chromatographic Assay Research Articles

Sensitive Ion-Chromatographic Determination of Citric Acid in Urine

Urine citrate analysis is relevant in the screening and monitoring of patients with calcium nephrolithiasis. A sensitive, fast, easy, and low-maintenance ion chromatographic (IC) method with conductivity detection for the analysis of urine citrate is developed and validated. Its application on true samples is also reported. Sample urine is diluted with a water solution containing internal standard (IS) before the chromatographic assay. The isocratic chromatographic run time is twenty-five minutes, using sodium hydroxide aqueous solution as the mobile phase. The method is fully validated as a quantitative method to objectively demonstrate its applicability for the intended use. The analytical response is linear in the 0.08–10.4 mmol/L concentration range. Precision and accuracy studies carried out on spiked urine and internal quality control samples reveal an imprecision CV% lower than 11% and an accuracy between 85 and 103%. The stability of citrate in urine samples is also evaluated. An easy, rapid, and low-maintenance, cost-effective IC method for urinary citrate determination is developed and validated. Internal standardization improves reliability and precision. The method has been currently used in our laboratory over recent years to analyze more than 1000 samples per year.

Ultrasound-Assisted Extraction of Nannochloropsis oculata with Ethanol and Betaine: 1,2-Propanediol Eutectic Solvent for Antioxidant Pigment-Rich Extracts Retaining Nutritious the Residual Biomass.

The aim of this study was the development of an efficient “green” extraction method of Nannochloropsis oculata to produce antioxidant extracts and nutritious residual biomass. Twenty-one extraction methods were evaluated by measuring the reactivity with the Folin–Ciocalteu reagent: ultrasonication or maceration at different temperatures with different organic solvents, extraction at different pH values, enzyme-assisted extraction, encapsulation with β-cyclodextrin, and the use of natural deep eutectic solvents. Ultrasound-assisted extraction with ethanol or betaine: 1,2-propanediol in a molar ratio of 2:5 (BP) had optimal extractive capacity. Both extracts were evaluated with antioxidant assays and the ethanol extract exhibited significantly higher (at least twofold) values. The determination of carotenoids by LC-MS and HPLC-DAD revealed the dominance of violaxanthin and antheraxanthin and their fourfold higher concentrations in the ethanol extract. The 1H-NMR characterization of the ethanol extract confirmed the results of the colorimetric and chromatographic assays. The microalgal biomass was characterized before and after the extraction in terms of humidity, ash, carbohydrates, proteins, chlorophyll-a, carotenoids, and lipids; the identity and content of the latter were determined with gas chromatography. BP caused a smaller depletion of the lipids from the biomass compared to ethanol, but proteins, carbohydrates, and ash were at a higher content in the biomass obtained after ethanol extraction, whereas the biomass was dry and easy to handle. Although further optimization may take place for the scale-up of those procedures, our study paves the way for a green strategy for the valorization of microalgae in cosmetics without generating waste, since the remaining biomass can be used for aquafeed.

Biomarker Assay Validation by Mass Spectrometry

Decades of discussion and publication have gone into the guidance from the scientific community and the regulatory agencies on the use and validation of pharmacokinetic and toxicokinetic assays by chromatographic and ligand binding assays for the measurement of drugs and metabolites. These assay validations are well described in the FDA Guidance on Bioanalytical Methods Validation (BMV, 2018). While the BMV included biomarker assay validation, the focus was on understanding the challenges posed in validating biomarker assays and the importance of having reliable biomarker assays when used for regulatory submissions, rather than definition of the appropriate experiments to be performed. Different from PK bioanalysis, analysis of biomarkers can be challenging due to the presence of target analyte(s) in the control matrices used for calibrator and quality control sample preparation, and greater difficulty in procuring appropriate reference standards representative of the endogenous molecule. Several papers have been published offering recommendations for biomarker assay validation. The situational nature of biomarker applications necessitates fit-for-purpose (FFP) assay validation. A unifying theme for FFP analysis is that method validation requirements be consistent with the proposed context of use (COU) for any given biomarker. This communication provides specific recommendations for biomarker assay validation (BAV) by LC-MS, for both small and large molecule biomarkers. The consensus recommendations include creation of a validation plan that contains definition of the COU of the assay, use of the PK assay validation elements that support the COU, and definition of assay validation elements adapted to fit biomarker assays and the acceptance criteria for both.

Development and Validation of New RP-HPLC Method for Estimation of Pramipexole Dihydrochloride in Bulk and Pharmaceutical Formulation

A novel high-performance liquid chromatographic assay method was developed and validated for the quantitative determination of the anti-Parkinson agent pramipexole dihydrochloride monohydrate in bulk and its tablet dosage form. In this perspective, the chromatographic separation was accomplished on Eclipse XDB-12 C18 (150 mm x 4.6 mm, 5 μm particle size) column using UV detection at 263 nm. The mobile phase consisted of distilled water: acetonitrile (10: 90 v/v), run at a flow rate of 1.0 mL/min with isocratic elution. The method was validated in accordance with ICH guidelines by evaluating the system suitability, linearity, limits of detection (LOD) and quantitation (LOQ), precision, accuracy, specificity, selectivity and short-term stability. Our findings revealed that retention time for pramipexole dihydrochloride was found to be 5.2 minutes. The linearity range was established between 6.25-225.0 μg/mL with a mean recovery of 101.26 % ± 0.56. The limits of detection and quantification were determined to be 4.18 μg/mL and 12.66 μg/mL, respectively, indicating that the method is very sensitive. Intra and inter-day precision were within acceptable limits (RSD

Population pharmacokinetics of ciprofloxacin in critically ill patients receiving extracorporeal membrane oxygenation (an ASAP ECMO study)

IntroductionThis study aimed to describe the pharmacokinetics (PK) of ciprofloxacin in critically ill patients receiving ECMO and recommend a dosing regimen that provides adequate drug exposure. MethodsSerial blood samples were taken from ECMO patients receiving ciprofloxacin. Total ciprofloxacin concentrations were measured by chromatographic assay and analysed using a population PK approach with Pmetrics®. Dosing simulations were performed to ascertain the probability of target attainment (PTA) represented by the area under the curve to minimum inhibitory concentration ratio (AUC0−24/MIC) ≥ 125. ResultsEight patients were enrolled, of which three received concurrent continuous venovenous haemodiafiltration (CVVHDF). Ciprofloxacin was best described in a two-compartment model with total body weight and creatinine clearance (CrCL) included as significant predictors of PK. Patients not requiring renal replacement therapy generated a mean clearance of 11.08 L/h while patients receiving CVVHDF had a mean clearance of 1.51 L/h. Central and peripheral volume of distribution was 77.31 L and 90.71 L, respectively. ECMO variables were not found to be significant predictors of ciprofloxacin PK. Dosing simulations reported that a 400 mg 8 -hly regimen achieved > 72% PTA in all simulated patients with CrCL of 30 mL/min, 50 mL/min and 100 mL/min and total body weights of 60 kg and 100 kg at a MIC of 0.5 mg/L. ConclusionOur study reports that established dosing recommendations for critically ill patients not on ECMO provides sufficient drug exposure for maximal ciprofloxacin activity for ECMO patients. In line with non-ECMO critically ill adult PK studies, higher doses and therapeutic drug monitoring may be required for critically ill adult patients on ECMO.

Phytochemical investigations and development of ethosomal gel with Brassica oleraceae L. (Brassicaceae) extract: An innovative nano approach towards cosmetic and pharmaceutical industry

The study was aimed to develop and characterize plant extract (Brassica oleraceae) loaded ethosomal gel for the valuation of cosmeceutical applications with a credence that botanical extracts have a greater capability to be employed in the formulation of advanced pharmaceuticals. Phytochemical profiling via FT-IR analysis, total bioactive contents of different polarity extracts (ethanol, dichloromethane, ethyl-acetate and chloroform) and quantification by using High Performance Liquid Chromatographic (HPLC) assay was carried out. Capabilities against oxidative stress were prodded through 2, 2-Diphenyl-1-picrylhydrazyl (DPPH), reduction potential analyses. Besides this, a correlation amongst total bioactive contents (TPC/TFC) and antioxidant potentials (biological activities) was studied by multi-variate statistical analysis. An augmented ethosomal preparation, after characterization and stability studies was incorporated to the gel matrix of Carbopol-940; Test/ EG (Ethosomal gel) with a Control/NEG (Non-ethosomal gel) formulation. Both formulations were then subjected to in-vitro characterization (organoleptic evaluation, pH, conductivity and rheology) of 90 days. By permeation studies flux (μg/cm2/hr); enhancement ratio was calculated and characterization of release data was done by using different kinetic models. The plant extracts revealed the presence of different functional groups along with 39.6 ± 0.5 mgGAE/g; 26.56 ± 0.4 mgQE/g phenolic and flavonoid contents, respectively; HPLC assay confirmed the existence of Quercetin, Gallic, Caffiec, Benzoic, Syringic and Sinapic Acids with highest antioxidant activity of 87.75% ± 1.18 in hydro-alcoholic fraction. Optimized ethosomal formulation exhibited entrapment efficiency of 68.57 ± 1.25, smallest vesicle size of 228.6 ± 1.17 nm with − 21.3 ± 0.15, zeta potential and a homogeneous size distribution of 0.233 ± 0.02. Significant results were obtained after in-vitro characterization. Both formulations followed Korsmeyer-peppas model with flux 35.499 and 21.731 μg/cm2/hr respectively and enhancement ratio 1.63. Thus, a stable ethosomal loaded gel of Brassica oleraceae extract was successfully developed and characterized for cosmeceutical potential.

Development and Validation of Fast and Robust Stability Indicating RP-HPLC Method for Simultaneous Estimation of Tolperisone Hydrochloride and Etoricoxib in Pharmaceutical Dosage Form

Objectives: A precise, accurate and selective stability-indicating reverse phase high performance liquid chromatographic assay method has been developed for the simultaneous quantitative determination of Tolperisone hydrochloride and Etoricoxib in tablets. Methods: The chromatographic separation of drugs was attained by using Eclipse plus C18 (150 mm × 4.6 mm, Agilent 5µm) column at room temperature. The composition of mobile phase was mixture of 0.035M triethylamine (pH 3.0 adjusted with orthophosphoric acid) and acetonitrile in ratio of 70:30 v/v and flow rate of mobile phase 1.0 ml/min with isocratic elution. The signal of eluents was observed at 290 nm by using diode array detector. Results: The retention time of Tolperisone hydrochloride and Etoricoxib were found to be 2.826 min and 7.566 min, respectively. The linearity ranges for both drugs were found to be 5-15 μg/ml and the percent recoveries were found to be 99.39% and 99.15% for Tolperisone hydrochloride and Etoricoxib respectively. Various forced degradation conditions, such as alkali hydrolysis, acid hydrolysis, thermal degradation, photolytic degradation, and oxidative degradation, were applied to the drug ingredients and drug formulation. The degradants were efficiently separated from the drugs by using optimized chromatographic conditions. The developed method was validated as per recommendation parameters of International council on harmonization guidelines Q2(R1). Conclusion: The validation parameters indicate that the drug substances were efficiently separated from its degradants and developed method can be routinely applied for the simultaneous quantitative determination of Tolperisone hydrochloride and Etoricoxib in combined tablet formulation in the quality control laboratory.

Population Pharmacokinetics and Dosing Simulations of Ceftriaxone in Critically Ill Patients Receiving Extracorporeal Membrane Oxygenation (An ASAP ECMO Study).

BackgroundDespite the surge in use of extracorporeal membrane oxygenation (ECMO) in the adult intensive care unit, little guidance is available on the appropriate dosing of antimicrobials in this setting. Ceftriaxone is an antimicrobial with a high affinity to plasma protein, a property identified in the literature as susceptible to sequestration into extracorporeal circuits and hypothesised to require dosage adjustments in this setting.ObjectiveThe aim of this study was to describe the pharmacokinetics of ceftriaxone and identify the best dosing regimen for critically ill adult patients receiving ECMO.MethodsSerial blood samples were taken from patients receiving both ECMO and ceftriaxone. Total and unbound drug concentrations were measured in plasma by chromatographic assay and analysed using a population pharmacokinetic approach with Pmetrics®. Dosing simulations were performed to identify the optimal dosing strategy: 60 and 100% of time with free (unbound) drug concentration exceeding the minimum inhibitory concentration (fT>MIC).ResultsIn total, 14 patients were enrolled, of which three were receiving renal replacement therapy (RRT). Total and unbound ceftriaxone was best described in a two-compartment model with total body weight, serum albumin concentrations, creatinine clearance (CrCL), and the presence of RRT included as significant predictors of pharmacokinetics. Patients not on RRT generated a mean renal clearance of 0.90 L/h, non-renal clearance of 0.33 L/h, and central volume of distribution of 7.94 L. Patients on RRT exhibited a mean total clearance of 1.18 L/h. ECMO variables were not significant predictors of ceftriaxone pharmacokinetics. Steady-state dosing simulations found that dosages of 1 g every 12 h and 2 g every 24 h achieved >90% probabilities of target attainment in patients with CrCL of 0 mL/min with RRT and 30 and 100 mL/min and various serum albumin concentrations (17 and 26 g/L).ConclusionsDosing recommendations for critically ill adult patients not on ECMO appear to be sufficient for patients on ECMO. Patients exhibiting augmented renal clearance (> 130 mL/min) or treatment of less susceptible pathogens may require higher doses, which requires further investigation.Supplementary InformationThe online version contains supplementary material available at 10.1007/s40262-021-01106-x.

Validation and application of a simple and rapid stability-indicating liquid chromatographic assay for the quantification of caffeine from human saliva

A clinical trial is currently underway to examine the efficacy of using caffeine citrate to prevent intermittent hypoxemia in late preterm neonates. Determining caffeine concentration using saliva in this population would be preferable as it is less invasive than plasma sampling, but a suitable method of analysis is required. This paper presents the development and validation of a rapid, efficient and reproducible stability-indicating high-performance liquid chromatography (HPLC) method and extraction protocol for the quantification of caffeine present in saliva. The stability of extemporaneously prepared caffeine citrate solutions (at 20–25 °C) was determined, along with the stability of caffeine spiked saliva samples (at 20–25 and 2–8 °C), to ensure the suitability of the developed method in the analysis of clinical trial samples. Protein precipitation using acetonitrile ensured the complete removal of salivary proteins and resulted in extraction recovery of ≥95% for all samples. The HPLC assay following extraction was linear (R 2>0.99) over the range 0.3–50 µg/mL (lower limit of quantification 0.3 µg/mL). The accuracy of the quality control samples was 94–100% and the relative standard deviation (RSD) was <7% for all samples. Caffeine-spiked saliva samples were stable for three freeze-and-thaw cycles pre-extraction and up to 72 hr post-extraction. The extraction and HPLC methods described were thus suitable for the analysis of clinical trial samples from the Latte Dosage Trial. All caffeine solutions were physically and chemically stable, with concentrations at the end of the three-month test period being within 4% of the baseline concentrations, indicating appropriateness for use as clinical trial medications.

Greenness estimation of chromatographic assay for the determination of anthracycline-based antitumor drug in bacterial ghost matrix of Salmonella typhimurium

The determination of an anthracycline-based antitumor drug, doxorubicin (DOX), in the bacterial ghost (BG) matrix of Salmonella typhimurium and commercial injectable dosage form was performed using a rapid, sensitive, and environmentally greener “high-performance liquid chromatography (HPLC)” assay. DOX was determined chromatographically on a Nucleodur (150 mm × 4.6 mm) RP C18 column with a particle size of 5 μm. The greener solvent system consisted of ethanol, methanol, and acetic acid (50:49:1 percent v/v/v), which was delivered with a flow rate of 1.0 mL/min. DOX was detected at a wavelength of 480 nm. With a determination coefficient value of 0.9992, the suggested HPLC assay was linear in the range of 1–100 μg/mL. In addition, the suggested HPLC assay for DOX analysis was rapid (Rt = 2.19 min), accurate (percent recoveries = 98.12–101.55), precise (percent CV = 0.87–1.97), and sensitive. The applicability of suggested HPLC technique was demonstrated by measuring DOX in S. typhimurium BG matrix and commercial injectable dosage form. DOX recovery in commercial injectable dosage form was calculated to be 101.02 percent. The amount of DOX in BG matrix was determined to be 48.51 μg/mL. Using the AGREE metric technique, the overall analytical GREEnness (AGREE) score for the proposed HPLC assay was calculated to be 0.79, indicating an outstanding greenness profile. These findings suggested that the proposed HPLC assay might be used to analyze DOX in BG matrices and commercial dosage forms on a regular basis.

Chemical Fingerprinting, Antioxidant, and Anti-Inflammatory Potential of Hydroethanolic Extract of Trigonella foenum-graecum.

In the current study, the antioxidant and anti-inflammatory potential of hydroethanolic extract of T. foenum-graecum seeds was evaluated. Phenolic profiling of T. foenum-graecum was conducted through high-performance liquid chromatography-photodiode array (HPLC-PDA) as well as through the mass spectrometry technique to characterize compounds responsible for bioactivity, which confirmed almost 18 compounds, 13 of which were quantified through a chromatographic assay. In vitro antioxidant analysis of the extract exhibited substantial antioxidant activities with the lowest IC50 value of both DPPH (2,2-diphenyl-1-picrylhydrazyl) and ABTS (2,2′-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid) inhibition assays. The extract was found to be non-toxic against human RBCs and murine macrophage RAW 264.7 cells. Moreover, the extract significantly (p < 0.001) reduced the lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-α), intrlukin-6 (IL-6), prostaglandin E2 (PGE2), and nitric oxide (NO) in RAW 264.7 cells in a concentration-dependent manner. The hydroethanolic extract of T. foenum-graecum exhibited considerable anti-inflammatory potential by decreasing the cellular infiltration to the inflammatory site in both carrageenan-induced peritonitis and an air pouch model of inflammation. Pretreatment with T. foenum-graecum extract caused significant improvement in antioxidants such as superoxide dismutase (SOD), CAT (catalase), malondialdehyde (MDA), and myeloperoxidase (MPO) against oxidative stress induced by carrageenan. Based on our results of in vivo and in vitro experimentation, we concluded that hydroethanolic extract of T. foenum-graecum is a potential source of phenolic compounds with antioxidant and anti-inflammatory potential.

The effect of Moringa oleifera polysaccharides on the regulation of glucocorticoid-induced femoral head necrosis: In vitro and in vivo

One of the common challenges in using glucocorticoid in the long term is the development of femoral head necrosis. To address this challenge, the use of glucocorticoid suppressors like plant polysaccharides has been considered. In this study, Moringa oleifera polysaccharide was isolated through hot water–ethanol precipitation method and purified by DEAE-Sepharose fast flow column. Then, they were characterized by FT-IR, NMR, methylation, and chromatography assays. The polysaccharide biocompatibility was investigated by MTT assay and its effect on osteoblasts was evaluated by controlling gene expression. Also, the effect of polysaccharide on dexamethasone-induced femoral head necrosis in rats was assessed by hydroxyproline, hexosamine and morphometric parameters. The results show that 2 Da molecular weight polysaccharide is mainly composed of Rha, Ara, Fru, Xyl, Man and Gal in the molar ratio of 1.7:2.1:3.4:5.9:5.8:1.3. Meanwhile, MTT results on osteoblasts cells showed polysaccharide biocompatibility, while significantly reducing the negative effects of glucocorticoid. Likewise, polysaccharide significantly reduced the levels of apoptosis and intracellular ROS of glucocorticoid-induced femoral necrosis. Moreover, the results of gene expression indicated a decrease in the expression of TNF-α and IL-6 genes using polysaccharide, which is very effective in preventing apoptotic activity. Also, Polysaccharide increased bone density, bone volume per tissue volume, trabecular thickness, and the hexosamine to hydroxyproline ratio in the rat serum in the presence of glucocorticoids, which are very effective in the process of femoral head necrosis. Furthermore, polysaccharide significantly increases the OCN, RUNX2 and COL-1 genes expression in cartilage tissue, which is in line with the result of morphometric parameters. Overall, this study suggests that the use of polysaccharide could result in the treatment of femoral head necrosis.

Characterization of Structural Hemoglobin Variants by Top-Down Mass Spectrometry and R Programming Tools for Rapid Identification.

Hemoglobinopathies are one of the most prevalent genetic disorders, affecting millions throughout the world. These are caused by pathogenic variants in genes that control the production of hemoglobin (Hb) subunits. As the number of known Hb variants has increased, it has become more challenging to obtain unambiguous results from routine chromatographic assays employed in the clinical laboratory. Top-down proteomic analysis of Hb by mass spectrometry is a definitive method to directly characterize the sequences of intact subunits. Here, we apply "chimeric ion loading" to characterize Hb β subunit variants. In this technique, product ions derived from complementary dissociation techniques are accumulated in a multipole storage device before delivery to a 21 T Fourier-transform ion cyclotron resonance mass spectrometer for simultaneous detection. To further improve the efficiency of identification of Hb variants and localization of the mutation site(s), we developed an R programming script, "Variants Identifier", to search top-down data against a database containing accurate intact mass differences and diagnostic ions from investigated Hb variants. A second R script, "PredictDiag", was developed and employed to determine relevant diagnostic ions for additional Hb variants with known sequences. These two R scripts were successfully applied to the identification of a Hb δ-β fusion protein and other Hb variants. The combination of chimeric ion loading and the above R scripts enables rapid and reliable interpretation of top-down mass spectrometry data, regardless of activation type, for Hb variant identification.

The Performance of Two Rapid Antigen Tests During Population-Level Screening for SARS-CoV-2 Infection.

Background: SARS-CoV-2 antigen assays offer a rapid mean to diagnose and isolate infected individuals. However, their utility in population-level screening is unknown.Objectives: The performance of two antigen tests in detecting SARS-CoV-2 was assessed among individuals randomly selected in the community.Study Design: A prospective study that performed head-to-head comparison of two SARS-CoV-2 antigen assays. Individuals were recruited during community SARS-CoV-2 screening over 10 working days. Demographic and clinical data were collected. Standard Q COVID-19 Ag test, a point-of-care chromatographic assay, was conducted immediately, and then the sample was transported to the virology laboratory to perform PCR and the LIAISON SARS-CoV-2 Ag chemiluminesence immunoassay.Results: respiratory samples from 991 individuals were collected, and 62 were positive by PCR. Inconclusive PCR results were observed in 19 samples and were excluded. The median age of participants was 40.2 years (IQR 32.3–47.8), and 932 (94%) were males. Most (77.4%) of infections were asymptomatic. The sensitivity and the specificity of the LIAISON assay were 43.3% (95%CI 30.6–56.8) and 99.9% (95%CI 99.3–100). The Standard Q assay had lower sensitivity (30.6%, 95%CI 19.6–43.7) but similar specificity (98.8%, 95%CI, 97.8–99.4). Similarly, the LIAISON assay had higher positive predictive value (96.3%, 95%CI 81–99.9% vs. 63.3%, 95%CI, 43.9–80.1%). Both assays performed better in symptomatic patients and among samples with a low-cycle threshold (Ct < 25).Conclusion: In our setting of random community surveillance, rapid antigen testing of nasopharyngeal swabs by either LIAISON SARS-CoV-2 Ag (DiaSorin) or Standard Q COVID-19 Ag (SD Biosensor) was less sensitive to detecting SARS-CoV-2 than the TaqPath COVID-19 RT-PCR.

Heavy metal contamination and antibiotic residues in poultry feed and meat in Bangladesh

Presence of harmful contaminants and residues in poultry feed and meat have serious public health consequence. This study was carried out to identify and quantify antibiotic residues, heavy metals and toxins in poultry feed and meat in the two selected poultry production belts of Bangladesh. A total of 94 broiler feed samples and 60 broiler meat samples were collected and tested by Thin Layer Chromatography (TLC), High Performance Liquid Chromatography (HPLC) and Enzyme-linked Immunosorbent Assay (ELISA) for identification and quantification of the parameters. Antibiotic residues were detected in 18.89% of the feed samples, whereas, there were no toxin (Aflatoxin) positive samples. Among the antibiotic positive samples, Oxytetracycline (OTC) was found predominant and detected in 12.22% cases. The mean concentrations of Cadmium (Cd), Lead (Pb), Chromium (Cr) were found as 0.04 mg/kg, 1.28 mg/kg and 2.55 mg/kg respectively in feed samples. In the case of meat samples, the mean concentration of OTC, Ciprofloxacin (CIP), and Tetracycline (TCL) residues were found 8.67 ppb, 7.18 ppb and 0.81 ppb accordingly. The highest mean concentration of Oxytetracycline (OTC) (10.15 ppb) was found in samples collected from local poultry sellers, whereas, the highest mean concentration of Tetracycline (TCL) (1.35 ppb) and Ciprofloxacin (CIP) (10.62 ppb) were observed in the samples obtained from local farm. The highest percentage of TCL and CIP (64% and 48% respectively) were found in samples collected from local farm. Chlortetracycline (CTC) was found predominant (70%) in samples collected from Contract farms. On the other hand, out of 60 meat samples, Cd and Cr were detected in only one meat sample with concentration of 56.41 mg/kg and 14.44 mg/kg respectively. Lead was not detected in any of the meat samples. Asian Australas. J. Food Saf. Secur. 2021, 5 (2), 71-78

HPLC with fluorescence detection for the bioanalysis and pharmacokinetic study of Doxorubicin and Prodigiosin loaded on eco-friendly casein nanomicelles in rat plasma

A rapid, efficient, and sensitive liquid chromatographic assay hyphenated to fluorometric detector (HPLC-FLD) was developed and validated for the determination of doxorubicin (DXR) and prodigiosin (PDG) in rat plasma. The sample pre-treatment involves a protein precipitation with acetonitrile with satisfying extraction efficiency (98% and 85% for DXR and PDG, respectively). The chromatographic separation was accomplished using stationary phase: Agilent Zorbax Eclipse plus-C18 analytical column (250 × 4.6 mm, 5 μm) and gradient eluting mobile phase of ammonium acetate (pH = 3), acetonitrile and methanol with programmed fluorescence detection. As the proposed method has been validated, it was subsequently implemented to evaluate DXR and PDG loaded on novel eco-friendly Casein nano drug delivery system after intravenous injection in healthy rats. A comparative pharmacokinetics’ study was carried out in rats for DXR in free form, DXR alone entrapped in the nanomicelle and DXR with PDG entrapped in the nano micelle. After testing the differences in pharmacokinetic parameters of the different formulations using ANOVA, the results showed insignificant differences among the tested parameters. This indicates that the presented nanomicelle delivery system has succeeded to incorporate PDG and DXR in a hydrophilic, safe, and potent formulation. This novel nanomicelle has negligible effect on the distribution and elimination of DXR.

Biomarkers of DNA Oxidation Products: Links to Exposure and Disease in Public Health Studies.

Environmental exposure can increase the production of reactive oxygen species and deplete cellular antioxidants in humans, resulting in oxidatively generated damage to DNA that is both a useful biomarker of oxidative stress and indicator of carcinogenic hazard. Methods of oxidatively damaged DNA analysis have been developed and used in public health research since the 1990s. Advanced techniques detect specific lesions, but they might not be applicable to complex matrixes (e.g., tissues), small sample volume, and large-scale studies. The most reliable methods are characterized by (1) detecting relevant DNA oxidation products (e.g., premutagenic lesions), (2) not harboring technical problems, (3) being applicable to complex biological mixtures, and (4) having the ability to process a large number of samples in a reasonable period of time. Most effort has been devoted to the measurements of 8-oxo-7,8-dihydro-2'-deoxyguanine (8-oxodG), which can be analyzed by chromatographic, enzymic, and antibody-based methods. Results from validation trials have shown that certain chromatographic and enzymic assays (namely the comet assay) are superior techniques. The enzyme-modified comet assay has been popular because it is technically simpler than chromatographic assays. It is widely used in public health studies on environmental exposures such as outdoor air pollution. Validated biomarker assays on oxidatively damaged DNA have been used to fill knowledge gaps between findings in prospective cohort studies and hazards from contemporary sources of air pollution exposures. Results from each of these research fields feed into public health research as approaches to conduct primary prevention of diseases caused by environmental or occupational agents.

A NOVEL VALIDATED HIGH-PERFORMANCE LIQUID CHROMATOGRAPHIC STABILITY-INDICATING ASSAY METHOD IN REVERSE PHASE FOR ESTIMATION OF HYDROXYPROGESTERONE CAPROATE IN AN INJECTABLE FORMULATION.

A high-performance liquid chromatographic stability-indicating assay method in reverse-phase has been developed and validated for the quantitative measurement of Hydroxyprogesterone corporate in injectable formulation prepared in castor oil base containing excipient benzyl benzoate as preservative. The reverse-phase method was developed by using Sunfire C18 column having dimensions (150 x 4.6 mm x 5μm) with a mobile phase[A] comprised of water, acetonitrile, and Methanol mixed in 40:30:30 v/v ratio and mobile phase [B] comprised of water, acetonitrile, and Methanol mixed in 20:40:40 v/v ratio pumped gradient program at a flow rate of 1.0 ml/min where the thermostat of the column oven controlled at 30°C and sample compartment at 5°C. The injection volume is set at 20μL. Hydroxyprogesterone corporate has a retention time of 10 minutes with UV detection at 240 nm. The detector was showing linear response in a range of 0.25 μg/ml to 150 μg/ml for Hydroxyprogesterone corporate. The guidelines of the International Conference on Harmonization on guidance for industry (2005) validation of analytical procedures, text, and methodology Q2(R1) were followed for the validation of this novel method. For quantification of Hydroxyprogesterone corporate the developed method was found to be genuinely exact precise, accurate, linear, robust, rugged, stable. and cost-effective.

SORPTION OF INDAZIFLAM IN BRAZILIAN SOILS WITH DIFFERENT pH VALUES

ABSTRACT Pre-emergence herbicides, such as indaziflam, have agronomic efficiency when available in the soil solution, mainly in the superficial layer (0-10 cm), the region with the highest concentration of weed seed bank. However, information about the sorption of indaziflam in Brazilian soils still is scarce, since most studies available in the literature were conducted in soils of temperate regions. The objective of this study was to estimate the sorption of indaziflam using high-performance liquid chromatography (HPLC) and bioassay in a Latossolo Vermelho-Amarelo (Oxisol) and in a Cambissolo (Inceptisol), at pH values of 5.1 and 6.1. In the biossay, Sorghum bicolor was used as an indicator plant. Quantitative indaziflam data were obtained by HPLC, and the sorption and desorption coefficients were determined in the soils using Freundlich isotherms. Increasing the pH of the two soils from 5.1 to 6.1 resulted in reductions of C50 and sorption coefficient values. Desorption, a parameter estimated only by HPLC, showed an inverse behavior to that of sorption, that is, the lowest desorption was observed in the condition of highest sorption. It is concluded that the increase in pH of the studied soils reduces indaziflam sorption, influencing the recommendations of this herbicide, from both the agronomic and the environmental points of view. When the goal is to verify the presence or absence of indaziflam in the studied soils, the bioassay technique has satisfactory efficiency. Chromatographic assay is necessary when quantifying the concentration of this herbicide in the soil.

Quality assessment of five pharmaceutical products of different brands marketed in Nigeria

Many pharmaceutical products have been reported to be substandard and counterfeit. About 15% of all drugs circulating in some parts of Africa and Asia are believed to be counterfeit, with figures rising to as high as 50%. This study was therefore carried out to assess the quality of five groups of drugs of different brands manufactured by pharmaceutical companies in Nigeria. Qualitative and quantitative analysis were carried out on the drugs using United States, British and Indian Pharmacopoeias. Qualitative analysis investigated includes: friability test, hardness test, disintegration test, dissolution test, pH, and weight uniformity for tablets; specific gravity, flow rate, pH and weight per ml of syrups. The quantitative analysis includes percentage purity and drug assay determination for different constituents using titrimetric, spectrophotometric and high performance liquid chromatographic (HPLC) methods. From the study, it was established that the Nigeria brands of pharmaceutical products sampled generally conformed to the established qualitative and quantitative specifications. There was however a short slight fall in the chromatographic assay for vitamin B6 in the Blood tonic from one company and similarly, the vitamin B1 in the Blood tonic of one company were at variance to those from other companies. The result for the statistical analysis shows that there is uniformity in weight of all tablets analysed and it complies with the specification. Standard pharmaceutical analysis must therefore be regularly carried out on pharmaceutical products before marketing so as to ensure good health, safety, elimination of sub-standard drugs and prevention of over- or under-dosage, which may further affect or cause damage to the health of the users. Consumer’s choice of generics should therefore be based on the analytical results (assay results, bio-pharmaceutical results, and statistical results) of the product and the projected bioequivalence.