Hydrolysis Conditions Research Articles

Optimisation of low temperature (8ᵒC) enzymatic hydrolysis of acid whey using design of experiments (DOE) for the generation of thermally stable whey protein hydrolysates

The impact of enzymatic hydrolysis at 8ᵒC of liquid whey protein concentrate (WPC35) derived from acid casein manufacture using Prolyve 1000®, a microbial protease preparation, under free-fall pH conditions on the physicochemical and the thermal stability properties of the resultant hydrolysates was investigated. The hydrolysis process was optimised using a design of experiments (DOE) approach. Eleven hydrolysates (H1–H11) were generated using 3 factors x 2 levels, i.e., enzyme:substrate (E:S) 0.25–1.00% (v/w), starting pH 7.5–9.0 and incubation time 10–96 h. Hydrolysate degree of hydrolysis (DH) values ranged from 0.75 (H10) to 4.74% (H6) which increased to 1.91 (H10) and 7.24% (H6) following subsequent thermal processing (mimicking the evaporation process) at 54ᵒC x 15 min and 64ᵒC x 10 min. The apparent viscosity (ηapp) of all hydrolysates was measured to assess their heat stability during heating at 85ᵒC for up to 20 min, this was shown to be E:S and pH-dependent. While the unhydrolysed samples formed a gel on heating at ∼74-80ᵒC. Following DOE analysis, it was found that E:S had a significant impact on the DH, the extent of intact protein degradation (%Deg) and the thermal stability. The %Deg following processing at 64ᵒC showed a strong correlation (R2 = 0.924; p < 0.001) with ηapp, and thus this parameter may be used to predict the thermal stability of the hydrolysate samples. The optimised hydrolysis conditions for the generation of heat stable WPC35 hydrolysates during 8°C incubation were achieved using an E:S = 0.625% at a starting pH = 8.65. Performance of protein hydrolysis reactions at low temperature has potential to enhance process sustainability.

Development and validation of stability indicating UPLC methods for related substances and assay analyses of ricobendazole hydrochloride

Ricobendazole hydrochloride is an active ingredient of a veterinary antiparasitic drug. The aim of this study was to investigate the degradation of ricobendazole hydrochloride under stress and stability testing conditions, for which we developed and validated the first stability indicating, specific, precise, accurate, and robust assay and related substances UPLC methods. The Acquity UPLC BEH C18 column was used for the related substances and assay analyses of ricobendazole hydrochloride, and the analyses were performed at 25 °C sample and 30 °C column temperatures with a 2 µL injection volume. In both methods, a mixture of water and methanol (60:40, v/v) was used as the diluent, mobile phase A was a phosphate buffer (50 mM potassium dihydrogen phosphate solution, pH 3.2 ± 0.05, adjusted with 10% o-phosphoric acid), and mobile phase B was a mixture of mobile phase A and acetonitrile (50:50, v/v). For the analysis of related substances, a gradient elution system was used at a flow rate of 0.4 mL/min for 35 min with a detection wavelength of 220 nm, while for the assay analysis; a gradient elution system was used at a flow rate of 0.3 mL/min for 15 min with a detection wavelength of 290 nm. The calibration curves showed excellent linearity with high R-squared (R2) values for each compound, ricobendazole (0.9998, 0.249 – 3.740 µg/mL), ricobendazole amine (0.9998, 0.255 – 3.819 µg/mL), albendazole (0.9998, 0.255 – 7.646 µg/mL), and albendazole sulfone (1.0000, 0.251 – 15.090 µg/mL) indicating a strong correlation between the concentrations of the compounds and their respective peak areas in the UPLC analysis. The method showed excellent accuracy with relative standard deviation values of less than 2.5%. The stress and photostability studies showed that ricobendazole hydrochloride was insensitive to daylight and UV radiation and showed significant degradation at elevated temperature (85 °C, 9 days) and under all hydrolysis and oxidation conditions. The major impurity was ricobendazole amine under thermal and hydrolysis conditions, while albendazole sulfone was the major oxidative impurity.

RP-HPLC Method Development, Validation Including Stability Indicating and Forced Degradation Studies for the Estimation of Pemetrexed in API and Pharmaceutical Dosage Form

Stability indicating analytical RP-HPLC (Reverse Phase- High performance Liquid Chromatography) method was developed for the analysis of the drug in the presence of its degradation products and subsequent validation of Pemetrexed that was conducted in API and pharmaceutical dosage form. Pemetrexed was subjected to different stress conditions as per International Conference on Harmonization (ICH) guidelines, stress conditions applied were including the effect of acid, base, oxidative, hydrolysis, thermal and photolytic degradation conditions. Chromatographic separation of API (Active Pharmaceutical ingredient) was achieved on Water’s HPLC instrument and ODS C18, 250x4.6 mm, 5µm particle size column with gradient elution of 0.1% ortho phosphoric acid buffer: Acetonitrile 80:20 ratio was taken as the eluent (mobile phase). The buffer pH was maintained at 3, the flow rate was 1.0 ml/min and wavelength were detected at 225 nm. The Retention time (RT) of Pemetrexed was Found at 2.9 minutes, column temperature was kept ambient and the runtime was 6 min. System suitability parameters like theoretical plates, tailing factor, and Asymmetry values were showed at 3391, 1.41, and 1.101 respectively. The percentage drug purity was found to be 99.99%. Validation of the method was proved that linear relationship over the range of 25-150 µg/mL, with linear regression curve (correlation coefficient) r2 value was noticed at 0.999 and LOD (limit of detection), LOQ (limit of quantification) values was established at 0.33µg/mL, 0.99µg/mL respectively. Recovery studies carried out at the concentrations of 50%, 100% and 150% with percentage recovery values of 100.06, 99.8 and 99.87 respectively. The precision of intraday was 0.64% and inter-day was 0.95%. Robustness of flow rate and mobile phase was changed even though the method was Robust. Forced degradation studies were also conducted to check the stability and suitability of this method to resolve the degradation products. Method validation was proved to be accurate, precise, linear, repeatable and robust. Thus, the accepted method can be proceeded for the estimation of Pemetrexed in pharmaceutical dosage form in a daily basis, due to their simplicity, suitability, accuracy, robustness and reproducibility.

Sedimentary evolution of the western North Yellow Sea mud patch: A Holocene perspective

The western North Yellow Sea mud patch has a unique location, abundant sediment supply and a complex sedimentary environment. Reconstructing its sedimentary evolution is crucial for a comprehensive understanding of source to sink process in the North Yellow Sea. Using core W03 collected from the mud patch, this study unravels its Holocene history through analysis of sedimentology, mineralogy and micropalaeontology. The results indicate that core W03 can be divided into three depositional units (DU). DU3 (538-461 cm) is composed of earthy yellow clayey silt and characterized by a hard layer. It was formed in the near-coast environment between 10.3 and 9.8 cal kyr BP, a hiatus period between Melt Water Pulse 1B and 1C events. DU2 (460-341 cm) was formed during the period of 9.8–6.5 cal kyr BP. As sea level rose, the westward retreat of the Yellow River mouth caused the transportation of Yellow River-derived sediment over longer distances, leading to an increase in flaky minerals. The rising sea level also induced a shift towards a reducing environment in the western North Yellow Sea. DU1 (340-0 cm) was formed after 6.5 cal kyr BP. During this period, the study area remained in a shelf shallow sea environment characterized by relatively strong chemical hydrolysis and reducing conditions. The intensification of the Yellow Sea Warm Current and an increase in Yellow River sediment transported to the sea led to an obvious increase in sedimentation rate after 2.3 cal kyr BP.

Changes in various lignocellulose biomasses structure after microwave-assisted hydrotropic pretreatment

Effective pretreatment of lignocellulosic biomass combined with delignification is a condition for effective enzymatic hydrolysis of cellulose, which is of key significance for the efficiency of the process of production of cellulosic ethanol. Becoming aware of the full range of changes in the structure of lignocellulose as a result of biomass pretreatment is important in view of the necessity to optimise the process of technological production of bioethanol. Our research was aimed at a comprehensive analysis of changes in structure of pine chip (softwood), beech chip (hardwood) and wheat straw (non-wood) biomasses resulting from a newly developed pretreatment method combining the use of hydrotrope in the form of sodium cumene sulfonate and microwave radiation. The performed analyses of the biomass composition indicate a high effectiveness of the proposed pretreatment method in the area of biomass delignification and hemicellulose removal. An effective removal of amorphous substances, i.e. lignin and hemicellulose, led to an increase in the crystallinity of pine chip biomass to 55%, and beech chips and wheat straw biomasses to 61%.

Isolation and Structural Characterization of a major degradation product of Ambrisentan using Q-TOF-MS, NMR and FTIR

Ambrisentan, an endothelin receptor antagonist, is subjected to forced degradation under neutral hydrolytic conditions, leading to a potential unknown impurity. This neutral degradation product (DP) has been detected using reverse-phase ultra-performance liquid chromatography (UPLC), where it has eluted at 1.24 relative retention time to the Ambrisentan peak. DP was isolated using semi-preparative HPLC from the degradation mixture. Later, the structure of DP was elucidated using high-resolution mass spectrometry, 2D Nuclear magnetic resonance spectroscopy (NMR), and Fourier transformed infrared spectroscopy (FTIR) spectroscopic techniques and characterized as 2-(2,2- diphenylvinyloxy)-4,6-dimethylpyrimidine.

Potato Peel Waste as an Economic Feedstock for PHA Production by Bacillus circulans.

Polymers of hydroxy alkanoates (PHA), also known as biodegradable, biocompatible plastic, are potential alternatives to petrochemical-based plastics. PHA is synthesized by microbes in their cytoplasm in the form of inclusion bodies in stress conditions such as nitrogen, oxygen, and phosphorus with excessive amounts of carbon. Sugar extracted from potato peel in the form of hydrolysate was employed as a carbon source for PHA production after acidic hydrolysis. The acid hydrolysis conditions are optimized for dilute acid concentrations and temperatures. The highest sugar-yielding condition (2% 15min at 121 ℃) was used for submerged fermentation for PHA production by Bacillus circulans MTCC 8167. Fourier transform infrared spectroscopy, nuclear magnetic resonance, and differential scanning calorimetry were used for polymer characterization. Gas chromatography coupled with mass spectrometry confirmed the monomers such as hexadecenoic acid 3-hydroxy, methyl esters, pentadecanoic acid 14 methyl esters, and tetradecanoic acid 12- methyl esters. Crotonic acid assay was used for quantification of PHA and it was found highest (0.232 ± 0.04g/L) at 37°C and 36h of incubation. Hence, potato peel waste could be a potential feedstock for waste to valuable production.

Evaluation of Interactions of Added Soybean Peroxidase with Other Nutrients Present in Fish Feeds Using an In Vitro Digestive Simulation.

Peroxidase (PO) has been applied in different areas of industrial biotechnology, including the control of contaminants like aflatoxin B1 in fish feeds. However, its potential negative interactions with the macro and micro components of feeds have not been evaluated. The aim of this study was to evaluate the impact of PO's addition to a feed on compounds like fatty acids and polyphenols using an in vitro simulation of the digestive tract of the tilapia. The influence on fatty acids was determined by changes in the peroxide index, with the feed including PO presenting values four times higher than those of the control feed. On the other hand, the in vitro digestive simulation also evidenced an effect of PO on the bioaccessibility of polyphenols significantly influenced by the total digestion time and temperature. The bioaccessibility of polyphenol ranged from 2.09 to 16.23 μmol of the total Trolox equivalent antioxidant capacity for the combinations evaluated in the study. The greatest bioaccessibility was observed at the central point under the following conditions of digestive hydrolysis: pH of 7, 30 °C, 4.5 h of digestive hydrolysis and an absence of PO.

Optimization of thermochemical hydrolysis of potassium hydroxide-delignified Cocoa (Theobroma cacao L) pod husks under low combined severity factors (CSF) conditions

This study identified the optimal conditions for acid hydrolysis of cocoa pod husks (CPH) pretreated and then hydrolyzed under combined low gravity conditions. Pre-treatment was carried out using 1% potassium hydroxide (KOH), using a biomass to solvent ratio of 9.143% (w/v), for 4 h at 30 °C, then the mixture is autoclaved for 15 min at 128 °C. Lignocellulosic analyzes were carried out on raw and pretreated husk. In order to determine the structural changes operated on the pretreated pod, Fourier transform infrared (FTIR) analyzes were also carried out. These analyzes confirmed a 93.87% removal of lignin after delignification. Furthermore, scanning electron microscopy/energy dispersive X‐ray spectrometry (SEM/EDX) revealed several changes within the pretreated matrix; showing a dominant presence of the elements carbon, oxygen and potassium, associated with an intensification of the peaks of magnesium and potassium. Acid hydrolysis on the pretreated pod was optimized under low combined severities factors (CSF) conditions between 1.11 and 2.45, using response surface methodology. A three-variable Box-Behnken design (autoclaving time (X1), biomass to solvent ratio (X2), and sulfuric acid (H2SO4) concentration (X3)) was used. The quadratic model with an R2 of 0.98 proved to be the most suitable, giving the following optimal hydrolysis conditions: 26.95 min of autoclave, biomass to solvent ratio of 9.53% (w/v) and 4.92% (w/v) H2SO4.18.77 g/L of reducing sugars was obtained for a CSF of 2.06. In addition, a marked decrease in the content of phenolic compounds was observed, shifting from 19.45 g/L in the raw delignified sample to 3.11 g/L in the delignified sample after reduction the extractable compounds. Thus, it is proved in this study that optimization of the acid hydrolysis process of CPH, after delignification with KOH, can improve the acid hydrolysis of polysaccharides, under conditions of low combined severity factors.

Açai Seeds (Euterpe oleracea Mart) Are Agroindustrial Waste with High Potential to Produce Low-Cost Substrates after Acid Hydrolysis.

Açai seeds have been discarded improperly around the Amazonia region, but they can be seen as promising low-cost substrates for fermentation processes. The structural carbohydrates and physicochemical characterization of açai seeds from the Amazonia were assessed followed by the determination of the optimal hydrolysis conditions using H3PO4 (phosphoric acid) and H2SO4 (sulfuric acid) to obtain a liquor with high contents of simple carbohydrates and low levels of potential microbial inhibitors usually generated during acid hydrolysis of carbohydrates. A central composite rotational design was carried out varying the concentrations of diluted acid (0-5%, w/v), solids (0.1-25%, w/v), and hydrolysis time (9.5-110 min). Acid hydrolysis with H2SO4 was more effective in producing reducing sugars (15.9-103.1 g/L) than H3PO4 (2.9-33.9 g/L) during optimization. The optimal hydrolysis conditions with H2SO4 were 3.5% of acid (w/v), 25% of solids during 70 min at 121 °C, which provided a liquor with 55 g/L of reducing sugars and low levels of microbial inhibitors: acetic acid (1.8 g/L), hydroxymethyl furfural (338 mg/L), and furfural (10 mg/L). Thus, açai seeds were characterized as promising agroindustrial waste with high potential to be used as a low-cost substrate in biotechnological processes, comprising relevant environmental and bioeconomic aspects for the development of the Amazonia.

Verification of Reinforced Surface Loose Layer of Zinc-Aluminum-Magnesium Steel Plate.

The corrosion resistance of zinc-aluminum-magnesium steel plates (Zn-Al-Mg steel plates) is significantly higher than that of galvanized steel plates. However, the unsatisfactory bonding performance of Zn-Al-Mg steel plates significantly limits their widespread application. In this study, X-ray photoelectron spectroscopy is employed to detect changes in the surface oxygen content of Zn-Al-Mg steel plates after different temperature treatments to confirm the existence of surface loose layers. In particular, changes in the surface oxygen content of the Zn-Al-Mg steel plates after the oxide layer is removed are investigated under saturated H2O vapor and O2 environmental conditions, and the cause of the formation of loose surface layers is determined. The uneven distribution of elements on the surface of the Zn-Al-Mg steel plates is investigated with scanning electron microscopy and energy dispersive spectroscopy. Nuclear magnetic resonance is employed to determine the size of the network spatial structure formed by silane coupling agents under different hydrolysis conditions and to further investigate the bonding performance of hydrolysate-modified Zn-Al-Mg steel plates. Several typical automotive adhesives are utilized to compare and examine the changes in the tensile strength of the Zn-Al-Mg steel plate bonding before and after modification with the silane coupling agent and analyze the structural damage of the adhesive at the bonding interface. The results confirm that the silane coupling agent strengthens the loose layer on the surface of the Zn-Al-Mg steel plate.

LC-PDA-QTof-MS characterization of the stress degradation products of Zanubrutinib: A novel BTK inhibitor

Zanubrutinib (ZAN) is an orally administered anti-cancer medication used for the treatment of Mantle cell lymphoma. Recently, it has also been approved by FDA for the treatment of chronic lymphocytic leukemia. Determination of impurities formed in drug substances/products as a result of manufacturing or storage forms an important aspect of drug life cycle management. The current study concentrated on understanding the stability of ZAN under various stress conditions as per the ICH Q1 (R2) guidelines. In total, ZAN produced thirteen degradation products under various hydrolytic (acid, base and neutral) and thermal stress conditions. The stress degradation products were separated by ultra-performance liquid chromatography, chemical structures of these products were characterized by MS/MS experiments combined with accurate mass measurements conducted on a LC-QTof-MS. The mechanism for the formation of these degradation products was also proposed. This study provides comprehensive information on the inherent stability of ZAN which will be useful in the drug development and manufacturing processes.

Study of hydrolysis and production of instant ginger (Zingiber officinale) tea

Abstract The aim of this study is to determine the suitable conditions for enzyme-assisted hydrolysis and the production of instant tea from hydrolysed ginger. Several parameters of ginger hydrolysis were investigated, such as the enzyme concentration from 0.1 to 0.4 g/100 g ginger powder (w/w), material/water ratios of 1/10; 1/15; 1/20; and 1/25 (w/w), pH from 3.5 to 5.0; temperature of hydrolysis process from 45 to 60°C, and time of hydrolysis process from 90 to 150 min. Subsequently, the hydrolysed ginger was spray-dried to produce instant products with surveyed parameters, including the maltodextrin concentration from 15 to 35 g/100 g ginger extraction, feed flow rates from 120 to 600 mL/h, and the temperatures of spray drying from 120 to 150°C. The suitable parameters for enzyme-assisted hydrolysis and production of instant tea were determined, including enzyme concentration as 0.2 g/100 g ginger powder (w/w), material/water ratio as 1/10 (w/w), pH at 5.0, temperature process as 55°C, the time of hydrolysis process as 120 min, the maltodextrin concentration as 15 g/100 g extraction (w/w), feed flow rate as 240 mL h−1, and the inlet temperature as 140°C. Furthermore, the physicochemical properties of the product were identified with moisture content as 2.36 ± 0.09%, total phenolic content as 98.34 ± 0.59 mg gallic acid equivalent/g DW, and IC50 value as 1,082 µgAA/mL.

High-Performance Methylsilsesquioxane Aerogels: Hydrolysis Mechanisms and Maximizing Compression Properties.

Synthesis of methylsilsesquioxane aerogels by ambient pressure drying instead of supercritical drying has recently emerged as a major trend, but the issues of low mechanical strength and unstable performance still need to be resolved. This work reveals the microscopic formation mechanisms of gel skeleton based on the kinetic characteristics of methyltrimethoxysilane (MTMS) precursor hydrolysis and the associated sol-gel reactions. The effects of oxalic acid concentration (cOA) and hydrolysis time of MTMS solution (th) on the gelation time, morphologies, microstructures, chemical structure, and compression properties of the as-synthesized methylsilsesquioxane aerogels are investigated. The optimal cOA and th are 38.4 mmol/L and 120 min, respectively, endowing the methylsilsesquioxane aerogels with a compression strength of 0.170 MPa and a maximum compression strain of 61.2%. Precise control of the hydrolysis conditions ensures the formation of branched particle-to-particle networks, which is crucial for maximizing the compression properties of methylsilsesquioxane aerogels synthesized under industry-relevant conditions.

Optimization of Enzyme-assisted Extraction of Bioactive Peptides from Whiteleg Shrimp (Litopenaeus vannamei) Head Waste Using Box-Behnken Design

Whiteleg shrimp (Litopeneaus vannamei) processing lines discard huge quantities of head wastes, which are the protein-rich source. Enzymatic hydrolysis has been studied for decades to salvage discarded whiteleg shrimp head waste to produce high-added value products applicable to animal industry. Alcalase 2.5 L, which is widely applied on marine materials, was used to hydrolyze whiteleg shrimp head waste protein and then purified by cold absolute ethanol. Hydrolysis conditions including enzyme/substrate (E/S) ratio, incubation temperature and duration were preliminarily screened (E/S ratio 0–2.5%, temperature 50–70 °C, duration 30–90 min) before conducting optimization using Box-Behnken design. After optimization, collected protein hydrolysate reached an experimental yield of 61.64% on dry basis under the optimal conditions as treated by Alcalase 2.5 L at 0.9% E/S ratio and incubated at 62 °C for 65 min. Protein hydrolysate performed bioactivity including DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical scavenging activity (3.88 ± 0.10 mg Trolox equivalents g dry matter–1), reducing power (19.20 ± 0.52 mg Trolox equivalents g dry matter–1) and α-amylase inhibitory activity (10.74 ± 0.65% inhibition at 50 mg mL–1).

Functional Properties and Proximate Analysis of Fish Waste Protein Hydrolysate Processed Using Enzymes

A huge production of fish and their processing waste give rise to by-products comprises up to 70% depending on the species, size and processing method. The waste includes visceral parts, head, frames, bones, skin and cut-offs are rich source of protein with high functional properties. It is generally discarded which is a wastage of nutrient source and leading to environmental issues. Therefore, it was aimed to utilized the by-products for maximum recovery of nutrients by enzyme hydrolysis method for the preparation of protein hydrolysate with the used of papain and pepsin for digestion following different hydrolysis conditions. With the hydrolysis of papain enzyme (1 to 6%), the protein content of finfish waste protein hydrolysate ranges from 19.17% ± 0.06 to 73.14% ± 0.08 and that of shellfish waste protein hydrolysate prepared with 5,10 and 15% papain enzyme showed 26.73% ± 0.04 to 40.4% ± 0.5 which is comparatively low. Whereas the highest protein content was observed in 1% pepsin enzyme treated finfish waste protein hydrolysate with 80.55% ± 0.07. Besides, the hydrolysates were composed of 6.91% ± 0.05 to 10.46% ± 0.05 (moisture content), 0.6% ± 0.01 to 2.4% ± 0.01 (ash content) and 0.02% ± 0.005 to 0.09% ± 0.005 (fat content). The hydrolysates were highly soluble ranges from 72.73% ± 0.05 to 93.83% ± 0.1 which indicates development of small size hydrophilic with highly solvated polypeptide particles. A reduced phenomena of foaming capacity and stability were observed in shellfish waste protein hydrolysate in contrast with finfish waste protein hydrolysate. Similar pattern was also resulted in emulsifying stability index. Whereas the emulsifying activity index was in the range of 6.15 ± 0.03 to 9.85 ± 0.07 m2/g. The water holding capacity of finfish and shellfish waste protein hydrolysate ranges from 3.4 to 4.23 gm/gm hydrolysate and 1.53 to 1.63 gm/gm hydrolysate respectively which is resulted by the difference in molecular weight of the peptide. The hydrolysates extracted from different sample with different enzyme and concentration at varying conditions were more or less similar ranges from 3.7 to 4.1 gm/gm protein hydrolysate (oil holding capacity). Hence, high protein content with good functional properties of the protein hydrolysate prepared with the utilization of fish waste is a positive impact on the attempt made to recover nutrient by enzymatic hydrolysis.

Hesperidin: Enrichment, forced degradation, and structural elucidation of potential degradation products using spectral techniques.

Hesperidin (HES) is a well-known citrus bioflavonoid phyto-nutraceutical agent with polypharmacological properties. After 2019, HES was widely used for prophylaxis and COVID-19 treatment. Moreover, it is commonly prescribed for treating varicose veins and other diseases in routine clinical practice. Pharmaceutical impurities and degradation products (DP) impact the drug's quality and safety and thus its effectiveness. Therefore, forced degradation studies help study drug stability, degradation mechanisms, and their DPs. This study was performed because stress stability studies using detailed structural characterization of hesperidin are currently unavailable in the literature. In the HES enrichment method crude HES was converted to its pure form (98% purity) using column chromatography and then subjected to forced degradation under acid, base, and neutral hydrolyses followed by oxidative, reductive, photolytic, and thermal stress testing (International Conference on Harmonization guidelines). The stability-indicating analytical method (SIAM) was developed to determine DPs using reversed-phase high-performance liquid chromatography (C18 column with methanol and 0.1% v/v acetic acid in deionized water [70:30, v/v] at 284 nm). Further, structural characterization of DPs was performed using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) and nuclear magnetic resonance (NMR) spectroscopy. In addition, in silico toxicity predictions were performed using pKCSM and DataWarior freeware. HES was found to be susceptible to acidic and basic hydrolytic conditions and yielded three DPs in each, which were detected using designed SIAM. Of six DPs, three were pseudo-DPs (short lived), and the remaining were characterized using LC-MS/MS and NMR spectroscopy. The tentative mechanism of the formation of proposed DPs was explained. The proposed DPs were found inactive from in silico toxicity predictions. Hesperidin was labile under acidic and basic stress conditions. The potential DPs were characterized using LC-ESI-MS/MS and NMR spectral techniques. The proposed mechanism of formation was hypothesized. In addition, to identify and characterize the DPs, a SIAM, which has broad biomedical applications, was successfully developed.

Optimizing reactor structure to increase MgH2 dispersion and improve hydrolysis performance

Hydrogen has attracted wide attention due to its high calorific value, good combustion performance, non-toxic, and green characteristics. MgH2 has obvious advantages in hydrogen storage due to its high hydrogen storage density, mild hydrolysis conditions, rich raw materials, etc. However, poor hydrolysis kinetics limits its application in hydrogen storage. Different from research on MgH2 material modification, the physical method was employed to increase the MgH2 dispersion and improve the hydrolysis kinetics. Furthermore, hydrolysis reactors, including tubular, flaky and finned units, were designed and optimized. The results indicate that enhancing the dispersion of magnesium hydride can significantly improve the hydrolysis kinetics. The use of hydrolysis reactors with highly dispersed MgH2 can effectively enhance the controllability and continuity during the hydrolysis process. This can be attributed to the reduction in the surface coating of unreacted magnesium hydride by the generated magnesium hydroxide, as well as the improved transfer performance between the fresh hydrolysis solution and solid magnesium hydride. The findings of this study can further expand the range of reactor types for magnesium hydride hydrolysis and provide new insights for designing controllable and continuous hydrolysis reactors.

A critical assessment on stability behaviour of Vorinostat using LC-MS-QTOF with H/D exchange and NMR

Vorinostat is the first USFDA-approved HDAC inhibitor for the treatment of cutaneous t-cell lymphoma. Vorinostat was exposed to ICH-recommended hydrolytic (acid, base, and neutral), oxidative, thermal, and photolytic stress conditions to understand the degradation behaviour. A Stability indicating LC method was developed and validated for separating and identifying forced degradation products. Under different stress conditions, six degradants were identified and characterized by LC-HRMS, MS/MS, and hydrogen-deuterium exchange mass studies. Vorinostat was found to be highly susceptible to the acidic and basic environment. In contrast, the drug substance was stable in the solid state under thermal and photolytic conditions whereas, it was found moderately stable when photolytic stress was provided to dissolved state of Vorinostat in acetonitrile-water. The degradants were identified as 7-amino-N-phenylheptanamide, 8-hydrazineyl-8-oxo-N-phenyloctanamide, 8-oxo-8-(phenylamino)octanoic acid, 8-oxo-8-(2-(7-oxo-7-(phenylamino)heptyl)hydrazineyl)-N-phenyloctanamide, 8,8′-(1-hydroxyhydrazine-1,2-diyl)bis(8-oxo-N-phenyloctanamide), and N1-((8-oxo-8-(phenylamino)octanoyl)oxy)-N8-phenyloctanediamide. The mechanistic explanation for the formation of each degradant in stability conditions has also been derived. The major degradants were also isolated/synthesized and characterized through 1H NMR for preparing impurity standards. Additionally, in-silico toxicity of the degradants was predicted in comparison to the drug, to identify whether any degradant has any specific type of toxicity and requires special focus to set specification limits during formulation development. The predicted toxicity indicated that the degradants have similar safety profile as that of the drug and specification can be set as per general impurity guideline.

An integrated strategy for rapid discovery and identification of the potential effective fragments of polysaccharides from Saposhnikoviae Radix

Ethnopharmacological relevanceSaposhnikoviae Radix (SR) is a traditional Chinese medicine, known as “Fangfeng”. As one of the main active components, Saposhnikoviae Radix polysaccharides (SP) demonstrated a range of biological activities, especially immunity regulation activity. Aims of the studyThis study aimed at exploring whether polysaccharides have activity after degradation, then discovering the potential effective fragments of SP. Materials and methodsHere we establish the chromatographic fingerprints method for 32 batches of 1-phenyl-3-methyl-5-pyrazolone (PMP) derivatives of oligosaccharides by HPLC, meanwhile evaluating its immunomodulatory activity in vivo. Then, the potential effective fragments of SP were screened out based on the spectrum-effect relationship analysis between fingerprints and the pharmacological results. Besides, liquid chromatography ion trap-time of flight mass spectrometry (LC-IT-TOF MS) coupled with multiple data-mining techniques was used to identify the potential effective oligosaccharides. ResultsThese findings showed that the hydrolysate of SP have significant immunomodulatory, and the immunity regulation activity varies under different hydrolysis conditions. The 4 potential effective peaks of the hydrolysate of SP were mined by spectrum-effect relationship. Finally, the chemical structure of 4 potential effective oligosaccharide fragments of SP was elucidated based on LC-IT-TOF MS. F10 was inferred tentatively to be Hex1→6Hex1→6Hex1→6Hex1→6Hex1→6Gal; F18 was confirmed to be Rhamnose; F14 was inferred tentatively to be Hex1→4Hex1→ 4Hex1→4Gal and F25 was tentatively inferred to be Ara1→6Gal. ConclusionsThis study may provide a sound experimental foundation in the exploration of the active fragments from macromolecular components with relatively complex structures such as polysaccharides.