An overview of genome engineering in plants, including its scope, technologies, progress and grand challenges.

Genome editing is a useful, adaptable, and favored technique for both functional genomics and crop enhancement. Over the years, rapidly evolving genome editing technologies, including clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR/Cas), transcription activator-like effector nucleases (TALENs), and zinc finger nucleases (ZFNs), have shown broad application prospects in gene function research and improvement of critical agronomic traits in many crops. These technologies have also opened up opportunities for plant breeding. These techniques provide excellent chances for the quick modification of crops and the advancement of plant science in the future. The current review describes various genome editing techniques and how they function, particularly CRISPR/Cas9 systems, which can contribute significantly to the most accurate characterization of genomic rearrangement and plant gene functions as well as the enhancement of critical traits in field crops. To accelerate the use of gene-editing technologies for crop enhancement, the speed editing strategy of gene-family members was designed. As it permits genome editing in numerous biological systems, the CRISPR technology provides a valuable edge in this regard that particularly captures the attention of scientists.

Biochemical and phenological characterization of diverse wheats and their association with drought tolerance genes.

Drought is one of the most important wheat production limiting factor, and can lead to severe yield losses. This study was designed to examine the effect of drought stress on wheat physiology and morphology under three different field capacities (FC) viz. 80% (control), 50% (moderate) and 30% (severe drought stress) in a diverse collection of wheat germplasm including cultivars, landraces, synthetic hexaploid and their derivatives. Traits like grain weight, thousand grain weight and biomass were reduced by 38.23%, 18.91% and 26.47% respectively at 30% FC, whereas the reduction rate for these traits at 50% FC were 19.57%, 8.88% and 18.68%. In principal component analysis (PCA), the first two components PC1 and PC2 accounted for 58.63% of the total variation and separated the cultivars and landraces from synthetic-based germplasm. Landraces showed wide range of phenotypic variations at 30% FC compared to synthetic-based germplasm and improved cultivars. However, least reduction in grain weight was observed in improved cultivars which indicated the progress in developing drought resilient cultivars. Allelic variations of the drought-related genes including TaSnRK2.9-5A, TaLTPs-11, TaLTPs-12, TaSAP-7B-, TaPPH-13, Dreb-B1 and 1fehw3 were significantly associated with the phenological traits under drought stress in all 91 wheats including 40 landraces, 9 varieties, 34 synthetic hexaploids and 8 synthetic derivatives. The favorable haplotypes of 1fehw3, Dreb-B1, TaLTPs-11 and TaLTPs-12 increased grain weight, and biomass. Our results iterated the fact that landraces could be promising source to deploy drought adaptability in wheat breeding. The study further identified drought tolerant wheat genetic resources across various backgrounds and identified favourable haplotypes of water-saving genes which should be considered to develop drought tolerant varieties.

Organic amendments improve salinity-induced osmotic and oxidative stress tolerance in Okra (Abelmoschus esculentus (L.)Moench).

AIMS: Salinity adversely affects okra [Abelmoschus esculentus (L.) Moench] plants by inducing osmotic and oxidative stresses. This study was designed to enhance salinity-induced osmotic and oxidative stress tolerance in okra plants by applying organic amendments. METHODS: The effects of different organic amendments (municipal solid waste compost, farmyard manure (FYM) and press mud) on osmotic potential, water use efficiency, activities of antioxidant enzymes, total soluble sugar, total soluble proline, total soluble protein and malondialdehyde (MDA) contents of okra plants grown under saline conditions (50 mM sodium chloride) were evaluated in a pot experiment. The organic amendments were applied each at the rate of 5% and 10% per pot or in various combinations (compost + FYM, FYM + press mud and compost + press mud each at the rate of 2.5% and 5% per pot). RESULTS: As compared to control, high total soluble sugar (60.41), total soluble proline (33.88%) and MDA (51%) contents and increased activities of antioxidant enzymes [superoxide dismutase (83.54%), catalase (78.61%), peroxidase (53.57%] in salinity-stressed okra plants, were indicative of oxidative stress. Salinity significantly reduced the osmotic potential (41.78%) and water use efficiency (4.75%) of okra plants compared to control. Under saline conditions, 5% (farmyard manure + press mud) was the most effective treatment, which significantly improved osmotic potential (27.05%), total soluble sugar (4.20%), total soluble protein (73.62%) and total soluble proline (23.20%) contents and superoxide dismutase activity (32.41%), compared to saline soil. Application of 2.5% (FYM + press mud), 5% press mud, and 10% compost significantly reduced MDA content (27%) and improved activities of catalase (38.64%) and peroxidase (48.29%), respectively, compared to saline soil, thus facilitated to alleviate oxidative stress in okra plants. CONCLUSIONS: Using organic amendments (municipal solid waste compost, farmyard manure and press mud) was a cost-effective approach to improve salinity-induced osmotic and oxidative stress tolerance in okra plants.

Agitation role (Dissolved Oxygen) in production of laccase from newly identified Ganoderma multistipitatum sp. nov. and its effect on mycelium morphology.

BACKGROUND: Agitation speed influenced the production rate of laccase. Orbital speed not only influenced the enzyme production, but was also effective to dissolve the oxygen during growth of mycelium, spores, and chlamydospores. Shear effects of speed greatly influenced the morphology of mycelium. METHODS: Ganoderma multistipitatum was identified by ITS marker. Phylogenetic tree was constructed for species identification. Qualitatively by plate method contained guaiacol indicator, while quantitatively by submerged fermentation and Central Composite Design applied on agitation parameter for maximum laccase potential of this species. The effects of agitation speed on mycelium morphology were observed under compound and scanning electron microscope. RESULTS: Statistical optimization of agitation conditions were performed by using response surface methodology to enhance the production of laccase from Ganoderma multistipitatum sp. nov. Maximum laccase yield (19.44 × 105 ± 0.28 U/L) was obtained at 150 rpm grown culture, which was higher than predicted value of laccase production (19.18 × 105 U/L) under aerobic conditions (150 rpm). The 150 rpm provided the continuous flush of oxygen. The DO (dissolved oxygen) was maximum (65%) for "27 h" incubation at 150 rpm during laccase synthesis. The statistical value of laccase production was minimum under anaerobic or nearly static condition of 50 rpm. The predicted (12.78 × 105 U/L) and obtained (12.82 × 105 U/L) yield was low at 50 rpm. Optimization of orbital shaking for aeration conditions were performed by the use of "Response Surface Methodology". The submerged shaking flasks were utilized as a nutrients growth medium to maximize the production of laccase from G. multistipitatum. The minimum incubation time highly influenced the laccase yield from 7 to 15 days via utilization of less cost-effective medium under a promising and eco-friendly method. The morphological effects of rpm on mycelium were examined under compound and scanning electron microscopy. Higher rpm (200, 230) shear the mycelium, while 150 to 200 rpm exhibited smoother and highly dense branches of mycelia. CONCLUSION: The shear forces of 200 rpm caused the damages of mycelium and cells autolysis with less laccase production. This study concluded that 150 rpm saved the life of mycelium and enhanced the production rate of enzymes.

Foliar melatonin ameliorates drought-induced alterations in enzyme activities of sugar and nitrogen metabolisms in cotton leaves.

Sugar and nitrogen metabolisms help plants maintain cellular homeostasis, stress tolerance, and sustainable growth in drought conditions. Melatonin, a potent antioxidant and signaling molecule, appears to mitigate the negative impacts of drought on plants. This study aimed to investigate the potential role of foliar-applied melatonin in ameliorating drought-induced alterations in leaf sugar and nitrogen metabolisms' enzyme activities during cotton flowering and boll formation. To date, no study has examined drought-induced sugar and nitrogen metabolisms' enzyme activity changes in cotton treated with foliar melatonin. Drought levels (FC1 = 75 ± 5%, FC2 = 60 ± 5%, and FC3 = 45 ± 5%) were maintained between 3 and 35 days after flowering (DAF), and melatonin (M) concentrations (0, 25, 50, and 100 µmol L-1 ) were applied at 3 and 21 DAF in a completely randomized design. M100 concentrations at low FC levels significantly enhanced leaf sugar and N-metabolic enzyme activities, such as sucrose synthase (65.56%) and glutamine synthetase (55.24%), compared to plants not treated with melatonin; peaking between 7 and 21 DAF and declining gradually with crop growth. Moreover, the M100 concentrations at all FC levels, particularly FC3, significantly increased the relative expression of GhSusB, GhSusC, SPS1, and SPS3 genes, indicating that melatonin improves leaf sugar and N-metabolism enzymatic activities under drought stress. Therefore, applying M100 concentrations to cotton foliage under FC3 conditions during reproductive stages improves leaf water status, sugar, and N-metabolism enzyme activities, demonstrating melatonin's potent anti-stress, osmoregulatory, and growth-promoting properties in overcoming drought stress in cotton crops. Future research into the molecular mechanisms of melatonin-mediated sugar and nitrogen metabolism enzyme activities in cotton leaves may lead to biotechnological methods to improve drought resilience in cotton and other crops.

A Novel Based Synthesis of Silver/Silver Chloride Nanoparticles from Stachys emodi Efficiently Controls Erwinia carotovora, the Causal Agent of Blackleg and Soft Rot of Potato.

In recent years, the biological synthesis of silver nanoparticles has captured researchers' attention due to their unique chemical, physical and biological properties. In this study, we report an efficient, nonhazardous, and eco-friendly method for the production of antibacterial silver/silver chloride nanoparticles utilizing the leaf extract of Stachys emodi. The synthesis of se-Ag/AgClNPs was confirmed using UV-visible spectroscopy, DPPH free radical scavenging activity, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and X-ray diffraction (XRD). An intense peak absorbance was observed at 437 nm from the UV-visible analysis. The Stachys emodi extract showed the highest DPPH scavenging activity (89.4%). FTIR analysis detected various bands that indicated the presence of important functional groups. The SEM morphological study revealed spherical-shaped nanoparticles having a size ranging from 20 to 70 nm. The XRD pattern showed the formation of a spherical crystal of NPs. The antibacterial activity performed against Erwinia carotovora showed the maximum inhibition by centrifuged silver nanoparticles alone (se-Ag/AgClNPs) and in combination with leaf extract (se-Ag/AgClNPs + LE) and leaf extract (LE) of 98%, 93%, and 62% respectively. These findings suggested that biosynthesized NPs can be used to control plant pathogens effectively.

Identification and selection of suitable landfill sites using GIS-based multi-criteria decision analysis in the Peshawar District, Pakistan.

Managing municipal solid waste (MSW) is indispensable for public health and quality living standards in urban and rural environments. Suitable landfill site selection (LSS) requires the consideration of many diverse criteria. This study uses various environmental, social and economic criteria in a sophisticated multi-criteria decision analysis (MCDA) within the geographical information system (GIS) and aims to provide a scientific basis for the identification of suitable landfill sites which can efficiently be used for MSW in one of the most urbanized cities (Peshawar district) in Pakistan. The methodology involves pairwise comparisons of the selected and ranked criteria using the analytical hierarchy process (AHP). A weighted linear combination (WLC) was applied to generate the most suitable landfill site for MSW disposal within the study area. The results show that approximately 11.4% (14,485.15 ha) and 16.6% (21,211.95 ha) of the total study area are either very highly suitable or highly suitable, 20.8% (26,525.56 ha) area is unsuitable and the remaining areas are either low (27.3%; 34,834.84 ha) or moderately suitable (23.8%; 30,356.84 ha) for the construction of landfill sites. It is recommended that using the proposed landfill sites would have the least consequences of public health risks and environmental degradation.

Pattern of dyslipidemia and associated factors in coronary artery disease patients in Khyber Pakhtunkhwa: A cross-sectional secondary data analysis.

Objectives: To assess the prevalence, pattern, and associated factors of dyslipidemia in patients with coronary artery disease (CAD) in the Northwest region of Pakistan. Method: A cross-sectional secondary data analysis was performed on CAD patients visiting cardiology clinics in selected hospitals from July to December 2019. A total of 362 patients were included via consecutive sampling. Dyslipidemia was operationalized according to the "National Cholesterol Education Program (NCEP ATP III) guidelines". Results: Mixed dyslipidemia was recorded in 92.26% of the patients, while isolated dyslipidemia was observed in 5.24%. A high prevalence of combined dyslipidemia with increased LDL-C, TG, and low HDL-C was noted. Contrarily, elevated LDL-C was the commonest single lipid disorder (84.25%). Hypercholesterolemia was the least common disorder. Increasing BMI was found to be independently associated with hypercholesterolemia (OR: 1.19). Similarly, age (OR: 0.97) and being a rural resident (OR: 2.61) were independent factors associated with hypertriglyceridemia. Furthermore, being an urban resident (OR: 2.25) and increasing BMI (OR: 1.77) were also significantly associated with high LDL-C. Conclusion: Mixed dyslipidemias were observed in the majority of the patients. Age, BMI, and residence were noted to be independently associated with abnormal lipids. Early screening and proper management should be encouraged to minimize this significant cardiovascular risk.

The potential role of dietary plant ingredients against mammary cancer: a comprehensive review.

Breast cancer is known as the most devastating cancer in the global female community and is considered as one of the severe health care burdens in both developed and developing countries. In many cases, breast cancer has shown resistance to chemotherapy, radiotherapy and hormonal therapy. Keeping in view these limitations, there is an urgent need to develop safe, readily available and effective breast anticancer treatments. Therefore, the scientists are keen in the extraction of plant-based phytochemicals (organosulfur compounds, betalains, capsaicinoids, terpenes, terpenoids, polyphenols, and flavonoids) and using them as breast anticancer agents. Results of numerous epidemiological investigations have revealed the promising role of phytochemicals in the prevention and treatment of breast cancer. The diverse classes of plant bioactive metabolites regulate different metabolic and molecular processes, which can delay the proliferation of cancers. These phytochemicals possess chemo-preventive properties as they down-regulate the expression of estrogen receptor-α, inhibit the proliferation of cancer cells, and cause cell cycle arrest by inducing apoptotic conditions in tumor cells. This review article discusses the potent role of various plant-based phytochemicals as potential therapeutic agents in the treatment or prevention of breast cancer along with the proposed mechanisms of action.

Hydroxyurea- a cost effective treatment in developing countries for Atypical Chronic Myeloid Leukemia (aCML): Case Report of Two Patients.

INTRODUCTION: Atypical chronic myeloid leukaemia (aCML) is a rare chronic myeloproliferative disorder with a poor prognosis. CASE REPORT: This case report presents two cases of male geriatric patients, both referred from primary care in rural areas and received at an urban clinic in a tertiary care hospital on separate instances. The first patient complained of low-grade fever (on/off), generalized body aches, rapid weight loss and shortness of breath for the last 2 months. The second patient arrived pale looking with symptoms of generalized body aches, dizziness and anorexia. Both patients were diagnosed to have aCML according to the World Health organization criteria. MANAGEMENT & OUTCOME: Both the patients were from a low economic bracket and were treated with Hydroxyurea a relatively economic medicine successfully. The follow-up lasted for 12 months in both cases. No progression to acute myeloid leukaemia (AML) or relapse was observed. DISCUSSION: This case report shows the promising results of Hydroxyurea in treating aCML and can be a cost effective alternate to other expensive treatments (allogeneic hematopoietic stem cell transplantation) and expensive medicines in lower and middle-income countries especially for resource-limited patients. These two cases show promising evidence for further studies to evaluate and conduct pharmaco-economic evaluations as well as clinical trials to compare hydroxyurea with other available alternative treatments for an affordable therapeutic option towards prevention of relapse and disease free survival after aCML.

Predictive factors and outcomes of severe community acquired pneumonia in patients with respiratory failure.

Objectives: To explore the risk factors, pathogens and outcomes of severe community-acquired pneumonia (SCAP) in patients with respiratory failure. Methods: A prospective observational study was conducted at Northwest General Hospital & Research Centre, Peshawar, Pakistan from February 2016 to October 2018. All patients with Community-acquired pneumonia (CAP) who fulfilled the inclusion criteria were recorded consecutively. Diagnosis of SCAP was made following the criteria established by the IDSA/ATS in the consensus guidelines on the management of CAP in adults published in 2007. In-hospital mortality was the main outcome. Results: The final analysis comprised a total of 100 patients with SCAP. The mean age was 60.0±18.01 years, and 54.0% were female patients. Afghani patients represented 22.0% of the total patients. The most common comorbidity associated with SCAP was hypertension (42.0%). The most commonly isolated etiological agents were Acinetobacter baumannii, followed by extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli. In-hospital mortality was 45%. On multivariate analysis, factors associated with in-hospital mortality were age (OR 1.054; 95%Cl 1.01-1.10; p=0.021), presence of two or more complications (OR 4.51; 95%Cl 1.18-17.28; p=0.028), septic shock (OR 6.44; 95%Cl 1.55-26.803; p=0.010), length of mechanical ventilation (OR 1.17; 95%Cl 1.01-1.40; p=0.043), and paO2 (OR 4.51; 95%Cl 1.18-17.28; p=0.004). Conclusion: A high mortality rate was observed in our study. Age, presence of two or more complications, septic shock, length of mechanical ventilation, and low paO2 were identified to be independent predictors of mortality for patients with SCAP.

Prevalence of depression and anxiety among general population in Pakistan during COVID-19 lockdown: An online-survey.

The present study's aim is to find the prevalence of two of the common indicators of mental health - depression and anxiety - and any correlation with socio-demographic indicators in the Pakistani population during the lockdown from 5 May to 25 July 2020. A cross-sectional survey was conducted using an online questionnaire sent to volunteer participants. A total of 1047 participants over 18 were recruited through convenience sampling. The survey targeted depression and anxiety levels, which were measured using a 14 item self-reporting Hospital Anxiety and Depression Scale (HADS). Out of the total sample population (N=354), 39.9% suffered from depression and 57.7% from anxiety. Binary logistical regressions indicated significant predictive associations of gender (OR=1.410), education (OR=9.311), residence (OR=0.370), household income (OR=0.579), previous psychiatric problems (OR=1.671), and previous psychiatric medication (OR=2.641). These were the key factors e associated with a significant increase in depression. Increases in anxiety levels were significantly linked to gender (OR=2.427), residence (OR=0.619), previous psychiatric problems (OR=1.166), and previous psychiatric medication (OR=7.330). These results suggest depression and anxiety were prevalent among the Pakistani population during the lockdown. Along with other measures to contain the spread of COVID-19, citizens' mental health needs the Pakistani government's urgent attention as well as that of mental health experts. Further large-scale, such as healthcare practitioners, should be undertaken to identify other mental health indicators that need to be monitored.

A clinical audit to evaluate antibiotic prescribing practice in pediatric patients admitted for enteric fever. Rationalizing antibiotic stewardship program.

To evaluate the antibiotic prescribing practice in pediatric patients for enteric fever, and to assess the need of developing and implementing the Antibiotic Stewardship Program (ASP) for the hospital. A prospective audit was completed in the pediatric ward of tertiary care hospital of Lahore for one year. Blood culture reports were collected from microbiology departed and clinical data were assessed regarding the choice of antibiotics, frequency, dosage and clinical outcome. All the statistics were analyzed using SPSS software and compared with the guidelines. Out of 157 cases hospitalized with suspicion of enteric fever, 137 cultures were positive for salmonella. Monotherapy of ceftriaxone (70%) was prescribed mostly as empirical therapy. About 20% of patients received a combination of antibiotics empirically. Susceptibility reports showed only 7 cases were of non-resistant typhoid, 15 multi-drug resistant and 115 extensively drug-resistant. Nearly 46% of patients were discharged earlier whose empirical therapy was changed either before or promptly after susceptibility reporting. Commonly used definitive antibiotics (32%) were a combination of azithromycin and meropenem. Inappropriate use of antibiotics was noted frequently as compared to the guidelines. However, recommendations themselves need to be reviewed as antibiotic resistance patterns are changing drastically.

Separation of constituents from Bergenia stracheyi (Hook. F. & Thoms.) Engl. by high-speed countercurrent chromatography with elution mode and its antidiabetic and antioxidant in vitro evaluation.

Diabetes, a metabolic disorder, is caused by a high blood sugar level. Diabetes is an increasing health issue and search for potent antidiabetic agents is desirable. Owing to its ethnomedicinal value, the Himalayan perennial herb Bergenia stracheyi (Hook. f. & Thoms.) Engl. (Saxifragaceae Juss) is used to treat diabetes. Herein, an efficient high-speed countercurrent chromatography with elution mode is reported for separation of active compounds from B. stracheyi. In current investigation, six main compounds including ß-arbutin (1), bergenin (2), 6-O-galloylarbutin (3), gallic acid (4), 11-O-galloylbergenin (5), and (-)-epicatechin 3-O-gallate (6) with above 95% purity were efficiently separated in a single run using biphasic tert-butyl methyl ether/n-butanol/methanol/water (1:3:1:5, v/v/v/v) solvent system. The structures of these compounds were characterized using spectral techniques and compared with the literature. Antidiabetic and antioxidant activities evaluation of the study samples showed that ß-arbutin (1) and 6-O-galloylarbutin (3) have a significant protective effect, especially at high dose against hydrogen peroxide induced oxidative injury. Our results might help further in-depth phytochemical and biological evaluation studies in search of potent antidiabetic compounds from B. stracheyi.

Separation and anti-inflammatory evaluation of phytochemical constituents from Pleurospermum candollei (Apiaceae) by high-speed countercurrent chromatography with continuous sample load.

Pleurospermum (Apiaceae) species possess a wide range of biological properties viz. analgesic, anti-inflammatory, antimalarial, and so on. Pleurospermum candollei (DC.) Benth. Ex C. B. Clark. is reported to cure diarrhea, gastric, respiratory, stomach, abdominal, joint, and back pain problems. In addition, it is also used for both male and female infertility. The present study deals with an efficient technique using high-speed countercurrent chromatography for separation of chemical components from the methanol extract of P. candollei. Notably, nine main compounds namely luteolin 7-O-glucoside (1), oxypeucedanin hydrate (2), pabulenol (3), bergapten (4), heptadecanoic acid (5), (E)-isoelemicin (6), trans-asarone (7), α-linolenic acid (8), and isoimperatorin (9) were very efficiently separated and isolated in pure form. Multiple injections were applied followed by two off-line recycling high-speed countercurrent chromatography. The inhibitory effect of nitric oxide production of all compounds was tested in the presence of 200 ng/mL lipopolysaccharide in RAW264.7 mice macrophage cells. The results demonstrated that compounds 7 and 8 effectively inhibited nitric oxide production, with IC50 values of 28.44 and 53.18 µM, respectively. This study thus validates the traditional claim of using P. candollei. Taken together, these findings will be useful in future research to find a potential candidate with anti-inflammatory properties.

Efficient extraction and purification of benzo[c]phenanthridine alkaloids from Macleaya cordata (Willd) R. Br. by combination of ultrahigh pressure extraction and pH-zone-refining counter-current chromatography with anti-breast cancer activity in vitro.

INTRODUCTION: Macleaya cordata (Willd) R. Br. (Papaveraceae family) is a well-known traditional Chinese medicine used to treat muscle pain, inflamed wounds, and bee bites. Benzo[c]phenanthridine alkaloids are the main active ingredients in M. cordata. In this work, sanguinarine and chelerythrine were efficiently extracted and purified by ultrahigh-pressure extraction (UHPE) technique and pH-zone-refining counter-current chromatography (PZRCCC) from M. cordata. OBJECTIVE: To develop an efficient UHPE method followed by an efficient separation technique using PZRCCC for benzo[c]phenanthridine alkaloids from the study plant species, and to evaluate the study samples for anti-breast cancer activity. METHODOLOGY: The optimal extraction conditions were optimised as extraction pressure 200 MPa, extraction solvent 95% ethanol, solid-liquid ratio 1:30 (g/mL) and extraction time 2 min. A two-phase n-hexane/ethyl acetate/i-propanol/water (1:3:1.5:4.5, v/v) solvent system was optimised with 10 mmol triethylamine in the upper phase and 10 mmol trifluoroacetic acid in lower phase in PZRCCC. The sample loading was optimised as 2.50 g. Moreover, the samples were evaluated for anti-breast cancer activity later on. RESULTS: The 2.50 g sample loading yielded 0.45 g of sanguinarine and 0.59 g chelerythrine in one-step separation using PZRCCC. The anti-breast cancer activities of sanguinarine and chelerythrine were found stronger than positive control (vincristine 5.04 µg/mL) with half-maximal inhibitory concentration values of 0.96 and 3.00 µg/mL, respectively. CONCLUSION: This study showed that the established methods were efficient in extraction (UHPE) and separation (PZRCCC) of the sanguinarine and chelerythrine from M. cordata.

A Simple and Efficient Two-Dimensional High-Speed Counter-Current Chromatography Linear Gradient and Isocratic Elution Modes for the Preparative Separation of Coumarins from Roots of Toddalia asiatica (Linn.) Lam.

Toddalia asiatica (L.) Lam. (Rutaceae) has shown a broad spectrum of biological properties, such as anti-inflammatory, antioxidant, antimicrobial, anti-HIV, and anticancer properties. The present study is concerned with the separation of the main components with broad partition coefficients (KD values) from T. asiatica, using linear gradient high-speed counter-current chromatography (LGCCC) combined with an off-line two-dimensional (2D) mode. Similar to the binary gradient HPLC, the LGCCC mode is operated by the adjustment of the proportion between the mobile phase of 5:5:1:9 (v/v) (pump A) and 5:5:4.5:5.5 (v/v) (pump B) in an n-hexane/ethyl acetate/methanol/water solvent system. The off-line 2D-CCC mode was used in this study for the secondary separation of two similar KD value compounds with n-hexane/ethyl acetate/methanol/water (5:5:4:6, v/v). Notably, six coumarins, namely, tomentin (1), toddalolactone (2), 5,7,8-trimethoxycoumarin (3), mexoticin (4), isopimpinellin (5), and toddanone (6), were efficiently separated. The structures of the pure compounds were elucidated by spectral techniques and compared with the literature.

Knowledge, Attitude, and Barriers Toward ADRs Reporting Among Health-Care Professionals at Tertiary Care Health Settings in Peshawar, Pakistan: A Web Based Study.

Background: Underreporting of adverse drug reactions (ADRs) is considered a major determinant of poor ADR signal detection in Pakistan. Considering this, the study was proposed to evaluate healthcare professionals' (HCPs) knowledge attitude toward and the barriers that discourse ADRs reporting. Methods: A cross-sectional survey was distributed among HCPs in 3 major tertiary care facilities of Peshawar. A self-administered, 31 items questionnaire was circulated online to collect the required information. Relative index ranking was used to identify the top barriers to the ADR reporting process. Results: HCPs (n = 322) were requested, and over one-third (n = 122) responded. Of the total, 97 (79.5%) were males, and by designation, 59(48.4%) were resident medical officers. About 45% of the HCPs did not identify the appropriate pharmacovigilance (PV) definition. More than half of the HCPs (52.2%) distinguished the appropriate PV purpose. Nearly 80% HCPs did not know the acceptable reporting time frame, while 22.1% HCPs knew that regulatory body for ADRs does not exist in Pakistan. The majority (95.08%) of the HCPs either strongly agreed or agreed that reporting an ADRs is a professional obligation and all the HCPs were of the opinion that PV should be taught in detail to HCPs. Exploring the barriers, it was identified that the key barriers to ADRs reporting were "unavailability of professional environment to discuss ADRs," Relative Importance Index (RII) = 0.813, "lack of incentives for reporting" (RII = 0.774), "lack of knowledge regarding reporting" (RII = 0.693), and "insufficient knowledge of pharmacotherapy in detecting ADRs" (RII = 0.662). In addition to these, "complicated reporting forms" (RII = 0.616), "lack of motivation for reporting ADRs" (RII = 0.610), and "absence of professional confidence" were seen as major hindrances in effective reporting of ADRs (RII = 0.598). Conclusion: Concerning PV and ADR reporting poor knowledge was noted. However, the majority of the HCPs showed an explicit attitude regarding ADRs reporting. The majority of the HCPs disclosed unavailability of professional environment to discuss about ADRs, lack of incentives, and how to report the main factors hindering the ADRs reporting. It is emphasized that health authorities carve out a niche for a well purposeful PV center and pledge educational activities and trainings for increasing understanding and approaches regarding reporting of ADR.

Cichorins D-F: Three New Compounds from Cichorium intybus and Their Biological Effects.

Cichorium intybus L., (chicory) is employed in various traditional medicines to treat a wide range of diseases and disorders. In the current investigation, two new naphthalane derivatives viz., cichorins D (1) and E (2), along with one new anthraquinone cichorin F (3), were isolated from Cichorium intybus. In addition, three previously reported compounds viz., ß-sitosterol (4), ß-sitosterol ß-glucopyranoside (5), and stigmasterol (6) were also isolated from Cichorium intybus. Their structures were established via extensive spectroscopic data, including 1D (1H and 13C) and 2D NMR (COSY, HSQC and HMBC), and ESIMS. Cichorin E (2) has a weak cytotoxic effect on breast cancer cells (MDA-MB-468: IC50: 85.9 µM) and Ewing's sarcoma cells (SK-N-MC: IC50: 71.1 µM); cichorin F (3) also illustrated weak cytotoxic effects on breast cancer cells (MDA-MB-468: IC50: 41.0 µM and MDA-MB-231: IC50: 45.6 µM), and SK-N-MC cells (IC50: 71.9 µM). Moreover compounds 1-3 did not show any promising anthelmintic effects.

Impact of methylglyoxal modification of chicken sarcoplasmic protein emulsions on emulsifying properties, rheological behavior and advanced glycation end products.

BACKGROUND: Protein modification is used to improve emulsion properties. However, there are limited reports on the effect of methylglyoxal (MGO) modification on emulsifying properties and on the production of advanced glycation end-products (AGEs) in chicken sarcoplasmic protein emulsion (SPE). In this study, MGO solution was dispersed into prepared emulsion (17 mg mL-1 sarcoplasmic-soybean oil (v/v 5:1)) at 0, 0.5, 5, 10, 30 and 50 mmol L-1 concentrations. Emulsifying activity index (EAI), emulsifying stability index (ESI), Z-average diameter, polydispersity index (PDI), zeta potential, rheological behavior (thermal condensation characteristics and fluidity) and AGEs in different concentrations of MGO SPE were measured. In addition, the effect of MGO on the levels of AGEs, lipid and protein oxidation of the emulsion as well as their relationship has also been analyzed. RESULTS: Our findings showed that ESI had the lowest value when MGO was added at a concentration of 10 mmol L-1 , while Z-average, PDI, carbonyl and AGEs had the highest value at the same concentration. Also, 10 mmol L-1 MGO played an important role in affecting the rheology of the emulsion. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results showed that the presence of myofibrillar proteins (MPs) in SPE was the main reason for the crosslinking of polymers which could be damaged by high concentration of MGO (>10 mmol L-1 ). CONCLUSION: Different concentration of MGO showed varying effects on emulsion properties and on the formation of AGEs in chicken SPE. Pearson's correlation analysis concluded that protein oxidation played a significant positive role during MGO modification. © 2020 Society of Chemical Industry.