High-performance Liquid Chromatography Technique Research Articles

Validation of An Efficient 2D-HPLC Method for The Determination of Pentazocine

Pentazocine is an opioid analgesic used to treat moderate to severe pains. The real analysis for the pharmaceutical products containing pentazocine is basically to ensure the correct dose and patient safety. This study developed and validated an improved high-performance liquid chromatography (HPLC) method in the selective and accurate quantification of pentazocine. Two-dimensional (2D) HPLC technique is employed to enhance the resolution and selectivity compared to conventional HPLC methods. The 2D-HPLC instrumentation consists of two C18 columns coupling with a switching valve to capture fractions from the first column, which is analyzed in the second column. The mobile phase was optimized to 45% acetonitrile and 55% water with 0.1% phosphoric acid. The method was validated in the International Conference on Harmonization guidelines and shows excellent linearity (R2=0.998), limit of detection of 1.58 μg/L, accuracy of 97.70–102.50%, and precision by relative standard deviation (RSD) of 1.02–4.20%. Selectivity was verified in resolving pentazocine from paracetamol, caffeine, ibuprofen, and oxycodone in laboratory mixtures. The utility of the 2D-HPLC method was demonstrated by accurate quantification of pentazocine in pharmaceutical injections and tablets unaffected in excipients. This research provides a rich validating technique to enhance the quality control testing of pharmaceuticals containing pentazocine.

Biosensor Strip for Rapid On-site Assessment of Levodopa Pharmacokinetics along with Motor Performance in Parkinson's Disease.

While levodopa (L-Dopa) is the primary treatment for alleviating Parkinson's disease (PD), its efficacy is hindered by challenges such as a short half-life and inconsistent plasma levels. As PD progresses, the rising need for increased and more frequent L-Dopa doses coupled with symptom fluctuations and dyskinesias underscores the urgency for improved comprehension of the interplay between L-Dopa levels and PD motor symptoms. Addressing this critical need, we present a decentralized testing method using a disposable biosensor strip and a universal slope (U-slope) calibration-free approach. This enables reliable, rapid, simple, and cost-effective decentralized L-Dopa measurements from capillary blood. A pilot study with PD persons demonstrates the ability to monitor real-time L-Dopa pharmacokinetics from fingerstick blood after oral L-Dopa-Carbidopa (C-Dopa) tablet administration. Correlating capillary blood L-Dopa levels with PD motor scores revealed a well-defined inverse correlation with temporal motor fluctuations. We compared the resulting dynamic capillary blood L-Dopa levels with plasma L-Dopa levels using the traditional but clinically impractical high-performance liquid chromatography technique. By providing timely feedback on a proper L-Dopa dosing regimen in a decentralized and rapid fashion, this new biosensing platform will facilitate tailored optimal L-Dopa dosing, towards improving symptom management and enhancing health-related quality of life.

Histidine-modified pillar[5]arene-functionalized mesoporous silica materials for highly selective enantioseparation

Chiral enantiomers, especially the enantiomers of chiral drugs often exhibit different pharmacological activity, metabolism and toxicity, thus it is of great research significance to scientifically and reasonably develop single chiral drugs with low toxicity and high efficiency. Among them, high performance liquid chromatographic techniques based on chiral stationary phases (CSPs) has become one of the most attractive methods used to evaluate the enantiomeric purity of single-enantiomers compound of pharmacological relevance. In this work, pillar[5]arene functionalized with L- and D-histidine, respectively, were modified on the surface of mesoporous silica as novel chiral stationary phases called L/DHis-BP5-Sil. Notably, L/D-histidine had the characteristics of low steric hindrance and easy derivatization. Although the π-π interaction of imidazole group was weaker than that of benzene ring, the benzene ring bonding imidazole-conjugated ring in the structure produced better enantioseparation effect. The results showed that L/DHis-BP5-Sil can separate a variety of complex structural enantiomers with excellent reproducibility, thermal stability and separation performance. Hence, the unique advantage of the highly selective separation of L/DHis-BP5-Sil provides new insights into the enantioseparation field.

One-Pot Interference-Based Colorimetric Detection of Melamine in Raw Milk via Green Tea-Modified Silver Nanostructures.

Detection of melamine has proven to be a challenge, requiring the use of complex analytical techniques. This study introduces an innovative, straightforward one-pot technique for qualitative assessment of the milk adulterant melamine. Originally, silver nanoparticles (AgNPs) were synthesized by utilizing green tea extract, which acted as both a reducing and sensing element. The synthesized AgNPs were characterized using UV-vis spectroscopy, X-ray diffraction, zeta potential, transmission electron microscopy, field emission scanning electron microscopy, and Fourier transform infrared spectroscopy. Melamine, rich in -NH2 groups, interacts with the biopolyphenols of green tea extract through hydrogen bonding. This interaction inhibits the formation of nanoparticles, resulting in a noticeable colorimetric response. The data obtained were confirmed by a standard UV-vis spectrophotometer and validated by the high-performance liquid chromatography technique. The limit of detection achieved by this scheme was quite low, falling below the permissible levels recommended by government bodies, e.g., the Food Safety and Standards Authority of India (FSSAI).

Unveiling carcinogenic pollutant levels in environmental water samples through facile fabrication of gold nanoparticles on sulfur-doped graphitic carbon nitride

Extensive utilization of pesticides and herbicides to boost agricultural production increased the environmental health risks, which can be mitigate with the aid of highly sensitive detection systems. In this study, an electrochemical sensor for monitoring the carcinogenic pesticides in the environmental samples has been developed based on sulfur-doped graphitic-carbon nitride–gold nanoparticles (SCN-AuNPs) nanohybrid. Thermal polycondensation of melamine with thiourea followed by solvent exfoliation via ultrasonication leads to SCN formation and electroless deposition of AuNPs on SCN leads to SCN-AuNPs nanohybrid synthesis. The chemical composition, S-doping, and the morphology of the nanohybrid were confirmed by various microscopic and spectroscopic tools. The as-synthesized nanohybrid was fabricated with glassy carbon (GC) electrode for determining the carcinogenic hydrazine (HZ) and atrazine (ATZ) in field water samples. The present sensor exhibited superior electrocatalytic activity than GC/SCN and GC/AuNPs electrodes due to the synergism between SCN and AuNPs and the amperometric studies showed the good linear range of detection of 20 nM–0.5 mM and 500 nM–0.5 mM with the limit of detection of 0.22 and 69 nM (S/N = 3) and excellent sensitivity of 1173.5 and 13.96 μA mM−1 cm−2 towards HZ and ATZ, respectively. Ultimately, the present sensor is exploited in environmental samples for monitoring HZ and ATZ and the obtained results are validated with high-performance liquid chromatography (HPLC) technique. The excellent recovery percentage and close agreement with the results of HPLC analysis proved the practicability of the present sensor. In addition, the as-prepared materials were utilized for the photocatalytic degradation of ATZ and the SCN-AuNPs nanohybrid exhibited higher photocatalytic activity with the removal efficiency of 93.6% at 90 min. Finally, the degradation mechanism was investigated and discussed.

Development and Validation of Fast and Sensitive RP-HPLC Stability-Indicating Method for Quantification of Piroxicam in Bulk Drug.

A simple, rapid, sensitive, and cost-effective green solvent-assisted reverse-phase high-performance liquid chromatographic technique, coupled with a photodiode array detector, was developed and validated for the estimation of piroxicam (PRXM). The chromatographic separation was achieved by using a C-18 (250 × 4.6) mm, 5-μm stationary phase and a mobile phase consisting of methanol and 0.1% ortho-phosphoric acid in water in a ratio of (80:20) v/v at a flow rate of 1ml/min. The detection was carried out at a wavelength of 254nm with a constant injection volume of 10μL throughout the analysis. The calibration curve was observed to be linear over the optimum concentration range of 50-300μgmL-1, with an R2 value of 0.9995. The developed method was validated as per the International Council for Harmonisation (ICH) Q2 (R1) guideline. Various parameters like selectivity/specificity, accuracy/recovery, linearity, precision, detection limit, quantitation limit, robustness and stability of analyte in solution were performed for the method validation. The PRXM was evaluated under stressed conditions, including acidic, basic, oxidative, thermal and photolytic, as per ICH Q1 (R2) guidelines. Significant degradation was observed in acidic and basic degradation conditions. Conversely, the drug substance showed stability when exposed to oxidative, photolytic and thermal degradation conditions.

Enhancing the Antibacterial Impact of Lipopeptide Extracted from Bacillus licheniformis as a Probiotic against MDR Acinetobacter baumannii.

The antibiotic resistance of microorganisms is escalating rapidly. Infections caused by opportunistic pathogens in immunocompromised individuals have prompted researchers to seek for potent and safe antibacterial agents. The purpose of this investigation was to explore the suppression of virulence gene expression, specifically the pga operon genes responsible in biofilm formation in Acinetobacter baumannii, through the utilization of metabolites obtained from probiotic bacteria. To assess the antimicrobial properties, standard strains of five probiotic bacteria were tested against a standard strain of multidrug-resistant (MDR) A. baumannii employing the agar gel diffusion technique. Following the identification of the most potent probiotic strain (Bacillus licheniformis), the existence of its LanA and LanM genes was confirmed using the polymerase chain reaction (PCR) test. High-performance liquid chromatography (HPLC) and fourier-transform infrared spectroscopy (FTIR) techniques were employed to identify the intended metabolite, which was found to be a lipopeptide nature. The minimum inhibitory concentration (MIC) values and anti-biofilm activity of the targeted metabolite were determined using a dilution method in 96-well microplates and field emission scanning electron microscopy (FE-SEM). Real-time PCR (qPCR) was utilized for comparing the expression of pga operon genes, including pgaABCD, in A. baumannii pre- and post-exposure to the derived lipopeptide. The MIC results indicated that the probiotic product inhibited the growth of A. baumannii at concentrations lower than those needed for conventional antibiotics. Furthermore, it was observed that the desired genes' expression decreased due to the effect of this substance. This research concludes that the B. licheniformis probiotic product could be a viable alternative for combating drug resistance in A. baumannii.

Rapid analysis of the chemical constituents of traditional Tibetan medicine Sbyor-bzo-ghi-wang using ballpoint electrospray ionization technique.

The chemical constituents of traditional Tibetan medicines (TTM) can be identified using high-performance liquid chromatography and high-resolution mass spectrometry (HPLC-MS/MS) technique. However, the HPLC-MS/MS technique requires the sample to be pretreated and then separated using the specific liquid chromatography method, which is time consuming. This study developed a ballpoint electrospray ionization (BPESI) technique for analyzing the chemical constituents of Sbyor-bzo-ghi-wang. This technique is a simple and inexpensive method for the rapid identification of the chemical constituents of TTMs. After the important parameters of the homemade BPESI device were optimized, the chemical constituents of Sbyor-bzo-ghi-wang were quickly identified without sample pretreatment. The raw data were converted to mzML file using MSConvert and then identified using SIRIUS 5 software. The results showed that 30 compounds were identified from Sbyor-bzo-ghi-wang, namely eight bile acids, six flavonoids, four alkaloids, three amino acids, and nine others. Compared to the ultra-high-performance liquid chromatography-Q/Orbitrap and high-resolution mass spectrometry (UHPLC-Q/Orbitrap HRMS) technique, the BPESI technique identified almost similar types of compounds and also a comparable number of compounds. Compared with the traditional HPLC-MS/MS methods, the BPESI technique does not require complex sample pretreatment and subsequent chromatographic separation steps; also it consumes a small quantity of samples. Therefore, BPESI can be used for the qualitative analysis of the chemical constituents of Sbyor-bzo-ghi-wang.

Nanostructured rhodamine B/aluminosilicate extracted sugarcane bagasse modified with tobacco-derived carbon quantum dot as ratiometric fluorescence probe for determination of tetracycline

Tetracycline (TC), as a widely used antibiotic, is very useful in treating bacterial infections. However, its residues in animal foodstuffs can enter the human body through the food cycle and causes severe and chronic diseases. On the other hand, due to its weak non-biodegradability, it is considered a threat to the environment. In this regard, the development of sensing methods to detect and measure TC is need of the hour. Herein, a dual-emission fluorescence sensor based on porous aluminosilicate structure (ASS) with rough surface hexagonal shape morphology and pore diameter less than 2 nm was prepared. The porous AAS was modified by post-modification method with blue carbon dots (CDT) and rhodamine B (RB) as two fluorophores to develop the ratiometric fluorescence (RF) sensor (CDT-AAS/RB). Nanostructured CDT-AAS/RB emitted two resolved peaks at 445 and 585 nm , which were dramatically quenched in the presence of TC. The RF sensor, with excellent sensitivity, was able to measure TC over the linear range of 0.001–150 μM with a limit of detection of 5.4 nM in the aqueous phosphate buffer. Moreover, the AAS component granted high selectivity and anti-interference ability to the sensor. In addition, the stability of the sensor was greatly improved due to the non-accumulation of CDT nanoparticles and RB molecules in the presence of the AAS. The proposed method was able to determine TC in complex real samples with satisfactory recovery, and the obtained results were validated with standard high-performance liquid chromatography technique.

Connecting Bisphenol A Exposure to PCOS: Findings from a Case-Control Investigation.

Polycystic Ovary Syndrome (PCOS) is a multifaceted condition influenced by genetic, hormonal, and environmental factors. Among environmental factors, Bisphenol A (BPA)-a recognized endocrine disruptor-has been implicated in the development of PCOS. The study aimed to compare BPA levels in women diagnosed with PCOS with those in healthy controls, using the high-performance liquid chromatography (HPLC) technique. The study involved 80 women diagnosed with PCOS and 50 healthy control participants. Demographic and biochemical parameters were recorded, including age, Body Mass Index (BMI), and levels of testosterone, estradiol, Luteinizing Hormone (LH), Follicle Stimulating Hormone (FSH), Prolactin (PRL), Dehydroepiandrosterone Sulfate (DHEA-S), Thyroid Stimulating Hormone (TSH), and Insulin Resistance as measured by the Homeostatic Model Assessment (HOMA-IR). Furthermore, BPA levels were measured using the HPLC technique. Women with PCOS exhibited significantly higher mean age and BMI compared to healthy controls (p = 0.01, p < 0.0001, respectively). Additionally, higher levels of testosterone (p = 0.04), LH (p = 0.03) and BPA (p < 0.0001) were observed in women with PCOS. However, estradiol, FSH, PRL, LH/FSH ratio, DHEA-S, and TSH levels were not significantly different between the two groups. HOMA-IR levels were not recorded for the control group. A notable positive relationship emerged between Bisphenol A and luteinizing hormone (LH) levels (r = 0.23, p = 0.03), also significant negative correlation appeared between Bisphenol A and thyroid-stimulating hormone (TSH) levels. This study found that women with PCOS have elevated BPA levels compared with healthy controls, showing a need for further research on the relationship between BPA exposure and the development of PCOS.

Free Isoflavone (Daidzein and Genistein) Content in Soybeans, Soybean Meal and Dried Soy Hypocotyl Sprout using High Performance Liquid Chromatography (HPLC)

Background: Phytoestrogens are natural plant glucosides found primarily in legumes and grains. Phytoestrogens can interact with oestrogen receptors in the body either by mimicking or blocking the effects of oestrogen. Isoflavones are a major subclass of phytoestrogens and often comprise a substantial part of livestock feeds. This study was focused on the detection and quantification of free isoflavones especially genistein and daidzein in soy-based ingredients (soybeans, soybean meal and dried soy hypocotyl sprout) which are incorporated in the animal feed formulations. Methods and Results: The simultaneous determination of daidzein and genistein in soybeans, soybean meal and dried soy hypocotyl sprout was carried out using the analytical technique high performance liquid chromatography (HPLC). The limits of detection and quantitation were 4.81 and 14.59 µg/100 g for daidzein and 3.06 and 9.58 µg/100 g for genistein respectively. The precision of the procedure was 2.73 per cent for daidzein and 2.88 per cent for genistein. The average daidzein and genistein concentration levels were (239.16 ± 0.21 and 238.68 ± 0.27 µg/100g) for soybeans, (397.24 ± 0.64 µg/100g and 443.85 ± 0.62) for soybean meal, (551.03 ± 0.54 and 742.59 ± 0.76 µg/100g) for dried soy hypocotyl sprout. The mean recovery percentages for daidzein and genistein were 92.8 per cent and 93.66 per cent respectively. The findings indicated that all the soy-derived components examined contained isoflavones and dried soy hypocotyl sprouts possessed higher concentrations of both daidzein and genistein.

Validation of Novel High-Performance Liquid Chromatography Method for Meropenem Quantification in Plasma

Eminent societies of clinicians worldwide advise the implementation of therapeutic monitoring of meropenem. Our aim was to validate the new high-performance liquid chromatography (HPLC) technique for the measurement of meropenem in plasma that we developed. The validation of the method was performed in accordance with the official European Medicines Agency (EMA) guideline for bioanalytical method validation through the assessment of the following validation parameters: linearity and limit of detection/quantification, trueness and precision, recovery, selectivity, matrix-effect assessment, dilution integrity, carry-over assessment, and stability. Our calibration curve was found to be linear over the concentration ranges of 1.25–100 μg/mL, which covers the therapeutic range of meropenem in patients. From the calibration curve, the limits of detection and quantification were calculated to be 0.4 μg/mL and 1.2 μg/mL, respectively. Recovery ranged from 81.7 to 95.9 percent. Intra-day truennes ranged from −1.9 to 2.6% and inter-day trueness ranged from −4.7 to 3.8%. Intra-day precision was less than 4 percent, while inter-day precision was less than 7 percent. Meropenem remained stable in the patient plasma sample for one week at −20°C. Our HPLC technique can be applied in clinical practice for the therapeutic monitoring of meropenem in critically ill and other vulnerable patients since it is simple, rapid, and reliable.

Turkish population-based screening for first identified changes of BRCA1 and BRCA2 genes in breast and/or ovarian cancer patients

IntroductionIt is known that BRCA1 and BRCA2 genes’ mutation carriers are predisposed to breast and ovarian cancers and other organ cancers such as prostate, colon and cervix. In the previous study performed at X University, all coding exons of both genes were screened by denaturing gradient gel electrophoresis (DGGE). In addition to various nonsense, missense mutations, polymorphisms and intronic region changes, seven novel missense mutations, including H513L, H816P and S1517Y in BRCA1 and S326R, G258P, E2903K and N2742S in BRCA2, had been identified.MethodsTo determine whether these unclassified variants are pathogenic, DNA samples of 150 healthy individuals without a known cancer history in the family were screened in this study for these seven novel missense mutations. These DNA samples were recruited from archives of previous polymorphism studies. PCR performed DNA amplifications, and denaturing high-performance liquid chromatography (DHPLC) techniques did mutation screenings.ResultsPeak patterns suggestive of a change in DNA fragments were considered for sequencing analyses. Analyses revealed that none of the 150 DNA samples had any change in the seven screened fragments. As a result, it is assumed that these seven mutations might be novel pathogenic mutations described in the Turkish population.ConclusionIn conclusion, these carriers must be informed about the mutation and given appropriate genetic counseling by their physicians. In addition, genetic testing must be offered to high-risk individuals (men/women) in the family so that it would be possible for other family members to have genetic counseling and contribute to disease prevention. On the other hand, these findings would contribute to current literature with novel results and shed light on future research.

New Stability Indicating HPLC Method for Related Substances in Dapagliflozin

The requirements established by the International Council for Harmonization (ICH) for the measurement of dapagliflozin from tablets were satisfied by the development and successful implementation of an reversed-phase high performance liquid chromatography (RP-HPLC) technique that is characterized by its speed, precision, and accuracy. In order to separate the samples, an isocratic mode was used on a Princeton C18 column that was operating at a flow rate of 1-mL per minute. The phase was composed of acetonitrile and 0.1% triethylamine in a volume-to-volume ratio of 50:50, with a pH of 5.0 and a detection wavelength of 224 nm. 5.163 minutes was the retention time that was determined to be best under the circumstances of the chromatographic analysis. Based on the correlation value of 0.999, it can be concluded that the procedure adhered to Beer-Lambert’s law within the measurement range of 10 to 70 μg/mL. The findings not only verified the claim that was made on the label, but they also validated the mean percent drug concentration that was specified for the commercial tablet formulation, which was 100.57%. Following the completion of an investigation, it was discovered that the approaches that were proposed have sufficient proportions of precision measurements, robustness, accuracy, range, and linearity. As part of the technique for stress testing, a number of criteria were evaluated and evaluated overall. The elements that were considered to be among these were light, humidity, temperature, oxidation, and pH (acid/base).

Novel HPLC Method Validation for Quantification of Dapagliflozin in Bulk Drug and Formulation

A new, steady, easy-to-use, and straightforward high-performance liquid chromatography (HPCL) technique was also tested to determine dapagliflozin concentration in medicinal and API forms. For the chromatographic extraction study, a Phenomenex Gemini-NX C18 Column was used. This column was filled with a mobile phase that was 70:30 v/v methanol to sodium 1-octanesulphonate. This experiment used a 20-liter syringe amount and a flow rate of 1-mL per minute. A bunch of factors were used to make sure the new method worked. The drug dapagliflozin was released at a wavelength of 203 nm after 6.5 ± 0.5 minutes. A correlation coefficient (r2) of 0.9994 between 15 to 55 μg/mL shows that the planned process is linear. The theoretical plates and peak tailing were found to be 4129 and 1.03, respectively. In terms of healing, the percent accuracy that was reached was within the acceptable range of 98 to 100. New suggestions say that the regular testing of dapagliflozin in large amounts must be done using a proven method that is unique and easy to repeat. Forced breakdown tests with drugs were used to show that the proposed method is stable. The product was tested according to the steps laid out in the ICH Q2R1 Guidelines. The test looked at many chemical processes, such as oxidation, photolysis, heat stress, and acids and alkaline hydrolysis.

Green Analytical HPLC Method Development of Acyclovir Loaded Floating Raft Formulation in Spiked Rabbit Plasma

Objective: The need of this investigation is to design eco-friendly, sensitive, and precise bioanalytical high-performance liquid chromatography (HPLC) techniques that leverage green analytical tools such as green analytical procedure index (GAPI), analytical eco scale (AES), and analytical greenness metric (AGREE) to analyze ACV-spiked rabbit plasma and ACV-loaded floating raft formulation (FRF) because The literature survey revealed insufficient data for the HPLC approach to FRF. Method: Methods involving plasma spikes mirroring the in-vivo models provide insights into potential real-life scenarios and the functionality of floating raft formulations loaded with ACV. The Design Expert tool optimized critical process parameters and identified significant factors influencing the chromatographic technique. Results: The CCD was used to optimize the flow rate (0.6 mL/min) and buffer solvent ratio (99:1) for separation. The C18 column (250 X 4.6 mm) was utilized to achieve the elution of ACV. The developed method was thoroughly validated through ICH guidelines. The ACV, lower limit of detection (LLoD), and lower limit of quantitation (LLoQ) retention times were established at 2.3 minutes, 15, and 20 ng/mL, respectively. Discussion: In summary, the study demonstrated the exceptional performance sensitivity, specificity, and robustness of the bioanalytical method. The method proved highly effective in quantifying ACV in plasma samples in terms of revalidatability, transferability, reproducibility, and the product development cycle. The developed method proves that using both design of experiments (DoE) and green analytical chemistry together creates a valid, eco-friendly, and reliable way to analyze the making of an ACV-loaded floating raft formulation.

Less is more: Validating a single method for comprehensive rh-insulin analysis

The objective of this current study is to establish a single method for potency and related proteins analysis of human insulin formulations using reverse-phase high performance liquid (RP-HPLC) chromatography technique which was validated and verified for the potency analysis in insulin formulations. Chromatographic separation was achieved using an octadecylsilane (C-18) stationary phase and a mobile phase composed of 55% (v/v) buffer (0.2 M sodium sulfate in water, {pH 2.3}) and 45% (v/v) acetonitrile. Detection was performed by UV detector at 214 nm with a flow rate of 1 ml/min and an injection volume of 20 µL, at 40°C. Currently there are separate methods available in Indian Pharmacopoeia for analysis of Potency and Related proteins in human insulin. We have validated a single method where quantitation of potency and related proteins can be performed in the same run. The method validation exhibited linearity over the concentration range of 0.08–4.5 mg/ml (r2=0.999) with limit of detection of 0.094 mg/ml The accuracy of the method was 99–102.8%. Thus, it is proposed that both potency and related proteins in insulin formulations can be precisely evaluated using a single run thus saving the time and cost for quality analysis of insulin preparations both at manufacturing and regulatory laboratories which in turn will increase the market availability of such standard quality insulin preparations for public health use.

Formulation and optimisation of Ozenoxacin topical nano-emulgel including a comprehensive methodology to qualify and validate the critical parameters of an in-vitro release test method and ex-vivo permeation test

Objective The purpose of this study was to formulate, optimize Ozenoxacin topical nano-emulsion using factorial design followed by to prepare and evaluate nano-emulgel using validated in-vitro release testing (IVRT) technique for determination of Ozenoxacin release rate along with ex-vivo permeation testing (EVPT). Significance: Nano-emulgel is a proven delivery system for poorly soluble substances works by enhancing the solubility and bioavailability. Factorial design provides a systematic and efficient means to study the effect of multiple factors on responses. IVRT is an USP compendia technique utilized for performance analysis of semi-solid formulations. Methods Nano-emulsion formulation optimization was done with factorial design, evaluated for globule size and % entrapment efficiency (EE). Nano-emulgels were characterized for assay, organic impurities, rheological behavior, IVRT, EVPT, and skin retention studies. IVRT validation was executed using vertical diffusion cells (VDCs). Results Ozenoxacin nano-emulsion was optimized with 1:1 ratio of Oil: Smix, 3:1 ratio of Surfactant:Co-Surfactant, and 15000 RPM of homogenization speed which resulted 414.6 ± 5.2 nm globule size and 92.8 ± 2.1% entrapment efficiency. Results confirmed that IVRT and Reversed Phase – High Performance Liquid Chromatographic techniques were validated as per regulatory guidelines. In-vitro, ex-vivo drug release, and skin retention from the optimized nano-emulgel formulation was comparatively higher (∼1.5 times) than that from the innovator (OZANEXTM) formulation. Conclusions Based on these results, Ozenoxacin nano-emulgel can be considered an effective alternative and was found to be stable at 40 °C/75% RH and 30 °C/75% RH storage condition for 6 months.

A comparative study on the nutritional composition, protein structure and effects on gut microbiota of 5 fermented soybean products (FSPs)

In this study, we conducted an analysis of the differences in nutrient composition and protein structure among various fermented soybean products and their impacts on the gut microbiota of rats. Conventional physicochemical analysis was employed to analyze the fundamental physicochemical composition of the samples. Additionally, we utilized high-performance liquid chromatography and ELISA techniques to quantify the presence of antinutritional compounds. Fourier infrared spectroscopy was applied to delineate the protein structure, while 16 s rRNA gene sequencing was conducted to evaluate alterations in gut microbiota abundance. Subsequently, KEGG was utilized for metabolic pathway analysis. Our findings revealed that fermented soybean products improved the nutritional profile of soybeans. Notably, Douchi exhibited the highest protein content at 52.18 g/100 g, denoting a 26.58 % increase, whereas natto showed a 24.98 % increase. Douchi and natto demonstrated the most substantial relative amino acid content, comprising 50.86 % and 49.04 % of the total samples, respectively. Moreover, the levels of antinutritional factors markedly decreased post-fermentation. Specifically, the α-helix content in doujiang decreased by 13.87 %, while the random coil content in soybean yogurt surged by 132.39 %. Rats that were fed FSP showcased notable enhancements in gut microbiota and associated metabolic pathways. A strong correlation was observed between nutrient composition, protein structure, and gut microbiota abundance. This study furnishes empirical evidence supporting the heightened nutritional attributes of FSPs.

A green perspective on simultaneous HPLC and UV spectrophotometric estimation of Udenafil and Dapoxetine hydrochloride in pharmaceutical formulations

Clinical research has affirmed that the combination of Udenafil and Dapoxetine hydrochloride yields greater improvement in erectile dysfunction compared to the use of individual medications. There is a requirement of precise quantitative method for accurate determination of Udenafil and Dapoxetine when taken together. Consequently, our interest lies in developing and validating analytical methods for the simultaneous estimation of Udenafil and Dapoxetine - Various strategies, such as UV spectrophotometric approaches and an innovative, straightforward, accurate, sensitive, and robust High-Performance Liquid Chromatography (HPLC) procedure, have been explored. An Isocratic HPLC approach was employed using reversed-phase CHEMSIL ODS C18 column (4.6 mm x 250 mm, 5 µm) as a stationary phase and detection wavelength (λ max) 230 nm for the two analytes in the chromatographic column. The mobile phase comprised of (70:30, v/v) mixture of Acetonitrile and 10 mM Ammonium formate buffer having pH 5, adjusted with formic acid. The elution was carried out at a constant flow rate of 1 mL/min. The UV simultaneous equation technique estimated the medicine's quantities and derived a simultaneous equation for DAP and UDN using 2 wavelengths, 230 nm and 298 nm. Furthermore, the two advanced methods underwent rigorous statistical validation, guided by the valuable ICH Q2R1 guidelines, affirming their effectiveness as a successful analytical tool for the concurrent analysis of both tablets in bulk and the synthetic mixture fabricated in the laboratory. To assess the environmental impact of contemporary UV spectrophotometric and HPLC techniques, the Green Analytical Greenness Metric (AGREE) software and the Green Analytical System Index (GAPI) were employed to evaluate the greenness profile. This examination confirmed the method's environmental friendliness concerning solvent usage, chemical ingredients, energy consumption, and waste production. The developed UV method is more sustainable and greener compare to HPLC method for simultaneous estimation of compound, which can be utilised by industry for quantification of each individual drug in their combination products.