Quantitative analysis of cephalexin in solid dosage form by Raman spectroscopy and chemometric tools.

OBJECTIVE: To use Raman Spectroscopy for qualitative and quantitative evaluation of pharmaceutical formulations of active pharmaceutical ingredient (API) of Cephalexin. SIGNIFICANCE: Raman Spectroscopy is a noninvasive, nondestructive, reliable and rapid detection technique used for various pharmaceutical drugs quantification. The present study explores the potential of Raman Spectroscopy for quantitative analysis of pharmaceutical drugs. METHOD: For qualitative and quantitative analysis of Cephalexin API, various standard samples containing less and more concentration of API than commercial tablet was prepared. To study spectral differences, the mean plot of all the samples was prepared. For qualitative analysis, Principal Component Analysis (PCA) and for quantitative analysis Partial Least Square Regression analysis (PLSR) was used. Both of these are Multivariate data analysis techniques and give reliable results as published in previous literature. RESULTS: PCA model distinguished all the Raman Spectral data related to the various Cephalexin solid dosage formulations whereas the PLSR model was used to calculate the concentration of different unknown formulations. For the PLSR model, RMSEC and RMSEP were determined to be 3.3953 and 3.8972, respectively. The prediction efficiency of this built PLSR model was found to be very good with a goodness of the model value (R2) of 0.98. The PLSR model also predicted the concentrations of Cephalexin formulations in the blind or unknown sample. CONCLUSION: These findings demonstrate that the Raman spectroscopy coupled to PLSR analysis could be regarded as a fast and effectively reliable tool for quantitative analysis of pharmaceutical drugs.

Co-application of copper nanoparticles and metal tolerant Bacillus sp. for improving growth of spinach plants in chromium contaminated soil.

Chromium (Cr) is classified as a toxic metal as it exerts harmful effects on plants and human life. Bacterial-assisted nano-phytoremediation is an emerging and environment friendly technique that can be used for the detoxification of such pollutants. In current study, pot experiment was conducted in which spinach plants were grown in soil containing chromium (0, 5, 10, 20 mgkg-1) and treated with selected strain of Bacillus sp. and Cu-O nanoparticle (CuONPs). Data related to plant's growth, physiological parameters, and biochemical tests was collected and analyzed using an appropriate statistical test. It was observed that under chromium stress, all plant's growth parameters were significantly enhanced in response to co-application of CuONPs and Bacillus sp. Similarly, higher levels of catalase, superoxide dismutase, malondialdehyde, and hydrogen peroxide were also observed. However, contents of anthocyanin, carotenoid, total chlorophyll, chlorophyll a & b, were lowered under chromium stress, which were raised in response to the combined application of CuONPs and Bacillus sp. Moreover, this co-application has significant positive effect on total soluble protein, free amino acid, and total phenolics. From this study, it was evident that combined application of Bacillus sp. and CuONP alleviated metal-induced toxicity in spinach plants. The findings from current study may provide new insights for agronomic research for the utilization of bacterial-assisted nano-phytoremediation of contaminated sites.

Targeted Analysis of Veterinary Drugs in Food Samples by Developing a High-Resolution Tandem Mass Spectral Library.

Veterinary drug residues present in foods can pose severe health threats to the population. The present study aims to develop a high-resolution mass spectral library of 158 veterinary drugs of 16 different classes for their rapid identification in food samples through liquid chromatography-high-resolution electrospray ionization-tandem mass spectrometry (LC-HR-ESI-MS/MS). Standard drugs were pooled according to their log P values and exact masses before analysis. Spectra were collected at system automated collision energy, i.e., of 25-60 eV and four predetermined collision energies (10, 20, 30, and 40 eV) for each compound using a schedule precursor list of [M + H]+, [M + Na]+, and [M + NH4]+ ions. The utility of the developed database was checked by analyzing food samples. A total of 17 veterinary drugs based on the reference standard retention times (RTs), HR-MS spectra, and MS/MS spectra were identified in the analyzed samples. Moreover, five veterinary drugs were selected for quantitative analysis, including doxycycline hyclate, lincomycin, sulfasalazine, moxifloxacin, and diphenoxylate, using liquid chromatography-ion trap mass-spectrometry (LC-IT-MS). Concentrations of the drug were obtained to vary from 0.0805 to 0.9731 mg/kg in food samples and were found to be exceeded in most of the cases as per the maximum residue levels described by Food and Agriculture Organization (FAO)/World Health Organization (WHO). The MS data were submitted to the MetaboLights online database (MTBLS2914). This study will help in the high-throughput screening of multiclass veterinary drugs in foodstuffs.

Simple two-step purification and characterisation of peroxidase from Citrullus colocynthis.

Peroxidase is a biotechnologically important enzyme. The purification of peroxidase from the root of Citrullus colocynthis was carried out in a simple two-step process with maximum purity level. The sample was extracted in a high salt buffer, and the enzyme was partially purified with a Q-Sepharose anion exchange column. Final purification was carried out with HighLoad 16/600 Superdex G-75 column. The purified protein was analysed with SDS gel electrophoresis, which suggested a single band of approximately 35 kDa. Further, the enzyme was identified with the help of Mass spectrometric analysis using an ESI-QTOF Mass spectrometer. The study will be helpful for the isolation and its commercial uses in biotechnology.

Acidotic and hypoxic tumor microenvironment induces changes to histone acetylation and methylation in oral squamous cell carcinoma.

Hypoxia and acidosis are ubiquitous hallmarks of the tumor microenvironment (TME), and in most solid cancers they have been linked to rewired cancer cell metabolism. These TME stresses are linked to changes in histone post-translational modifications (PTMs) such as methylation and acetylation, which lead to tumorigenesis and drug resistance. Hypoxic and acidotic TME cause changes in histone PTMs by impacting the activities of histone-modifying enzymes. These alterations are yet to be extensively explored in oral squamous cell carcinoma (OSCC), one of the most prevalent cancers in developing countries. Hypoxic, acidotic, and hypoxia with acidotic TME affecting histone acetylation and methylation in the CAL27 OSCC cell line was studied using LC-MS-based proteomics. The study identified several well-known histone marks, in the context of their functionality in gene regulation, such as H2AK9Ac, H3K36me3, and H4K16Ac. The results provide insights into the histone acetylation and methylation associated with hypoxic and acidotic TME, causing changes in their level in a position-dependent manner in the OSCC cell line. Hypoxia and acidosis, separately and in combination, cause differential impacts on histone methylation and acetylation in OSCC. The work will help uncover tumor cell adaptation to these stress stimuli in connection with histone crosstalk events.

PX-12 synergistically enhances the therapeutic efficacy of vorinostat under hypoxic tumor microenvironment in oral squamous cell carcinoma in vitro.

Hypoxia is a characteristic feature of solid tumors, including oral squamous cell carcinoma (OSCC), which causes therapeutic resistance. The hypoxia-inducible factor 1-alpha (HIF-1α) is a key regulator of hypoxic tumor microenvironment (TME) and a promising therapeutic target against solid tumors. Among other HIF-1α inhibitors, vorinostat (suberoylanilide hydroxamic acid, SAHA) is a histone deacetylase inhibitor (HDACi) targeting the stability of HIF-1α, and PX-12 (1-methylpropyl 2-imidazolyl disulfide) is a thioredoxin-1 (Trx-1) inhibitor preventing accumulation of HIF-1α. HDACis are effective against cancers; however, they are accompanied by several side effects along with an emerging resistance against it. This can be overcome by using HDACi in a combination regimen with Trx-1 inhibitor, as their inhibitory mechanisms are interconnected. HDACis inhibit Trx-1, leading to an increase in the production of reactive oxygen species (ROS) and inducing apoptosis in cancer cells; thus, the efficacy of HDACi can be elevated by using a Trx-1 inhibitor. In this study, we have tested the EC50 (half maximal effective concentration) doses of vorinostat and PX-12 on CAL-27 (an OSCC cell line) under both normoxic and hypoxic conditions. The combined EC50 dose of vorinostat and PX-12 is significantly reduced under hypoxia, and the interaction of PX-12 with vorinostat was evaluated by combination index (CI). An additive interaction between vorinostat and PX-12 was observed in normoxia, while a synergistic interaction was observed under hypoxia. This study provides the first evidence for vorinostat and PX-12 synergism under hypoxic TME, at the same time highlighting the therapeutically effective combination of vorinostat and PX-12 against OSCC in vitro.

Dysregulation of metalloproteins in ischemic heart disease patients with systolic dysfunction.

Ischemic heart disease (IHD) is the leading cause of mortality worldwide. Metalloproteins have been linked to human health and diseases. The molecular functions of metalloproteins in IHD is not well understood and require further exploration. The objective of this study was to find out the role of metalloproteins in the pericardial fluid of IHD patients having normal (EF > 45) and impaired (EF < 45) left ventricular ejection fraction (LVEF). IHD patients were grouped into two categories: LVEF<45 (n = 12) and LVEF >45 (n = 33). Pooled samples of pericardial fluid were fractionated by using ZOOM-isoelectric focusing (IEF) followed by further processing using one-dimensional gel electrophoresis (1D SDS-PAGE) and filter-aided sample preparation (FASP). Tryptic peptides of each fraction and differential bands were then analyzed by nano-LC-ESI-MS/MS. Protein identification was performed through a Mascot search engine using NCBI-Prot and SwissProt databases. A total of 1082 proteins including 154 metalloproteins were identified. In the differential bands, 60 metalloproteins were identified, while 115 metalloproteins were identified in all ZOOM-IEF fractions. Twelve differentially expressed metalloproteins were selected in the intense bands according to their molecular weight (MW) and isoelectric point (pI). The 12 differentially expressed metalloprotein includes ceruloplasmin, Prothrombin, Vitamin K-dependent protein, Fibulin-1, Ribosomal protein S6 kinase alpha-6, nidogen, partial, Serum albumin, Hemopexin, C-reactive protein, Serum amyloid P-component, and Intelectin-1 protein which were all up-regulated while serotransferrin is the only metalloprotein that was down-regulated in impaired (LVEF<45) group. Among the metalloproteins, Zn-binding proteins are 36.5 % followed by Ca-binging 32.2 %, and Fe-binging 12.2 %. KEGG, pathway analysis revealed the association of ceruloplasmin and serotransferrin with the ferroptosis pathway. In conclusion, 154 metalloproteins were identified of them the Zn-binding protein followed by Ca-binding and Fe-binding proteins were the most abundant metalloproteins. The two metalloproteins, the Cu-binding protein ceruloplasmin, and Fe-binding protein serotransferrin are involved in the ferroptosis pathway, an iron-dependent form of regulated cell death that has been linked to cardiac pathology, especially in IHD patients having impaired systolic (LVEF<45) dysfunction. However, further research is required to validate these findings.

Echinocystic Acid Bidesmoside Saponins from Microglossa afzelii O. Hoffm and Their Cytotoxic Activity against the CAL-27 Oral Squamous Carcinoma Cell Line.

This paper describes eight new triterpenoid saponins, including afzeliioside A (1), four acetylated afzeliiosides as pairs of inseparable regioisomers, called afzeliiosides B/C (2/3) and D/E (4/5), afzeliiosides F-H (6-8), and a known impatiprin C (9), which were isolated from the n-BuOH fraction of the liana of Microglossa afzelii. Their structures were established mainly by extensive spectroscopic analysis, including 1D and 2D NMR, HRFAB-MS, tandem ESI-MS/MS, and chemical methods, as well as a comparison of their spectral data with those of related compounds. All the isolates were screened for their cytotoxic activity against the CAL-27 oral squamous carcinoma cell line. Only compounds 4/5 (EC50 = 36.0 µg/mL (32.7 µM)) exhibited moderate cytotoxic activity. This work presents the first chemical and biological investigation of Microglossa afzelii and reports, for the first time, on the isolation of saponins in the genus Microglossa.

Sensitive Detection of Pharmaceutical Drugs and Metabolites in Serum Using Data-Independent Acquisition Mass Spectrometry and Open-Access Data Acquisition Tools.

Data-independent acquisition (DIA) based strategies have been explored in recent years for improving quantitative analysis of metabolites. However, the data analysis is challenging for DIA methods as the resulting spectra are highly multiplexed. Thus, the DIA mode requires advanced software analysis to facilitate the data deconvolution process. We proposed a pipeline for quantitative profiling of pharmaceutical drugs and serum metabolites in DIA mode after comparing the results obtained from full-scan, Data-dependent acquisition (DDA) and DIA modes. using open-access software. Pharmaceutical drugs (10) were pooled in healthy human serum and analysed by LC-ESI-QTOF-MS. MS1 full-scan and Data-dependent (MS2) results were used for identification using MS-DIAL software while deconvolution of MS1/MS2 spectra in DIA mode was achieved by using Skyline software. The results of acquisition methods for quantitative analysis validated the remarkable analytical performance of the constructed workflow, proving it to be a sensitive and reproducible pipeline for biological complex fluids.

Genomic characterization of high-risk Escherichia coli and Enterobacter hormaechei clones recovered from a single tertiary-care hospital in Pakistan.

AIMS: Spread of carbapenem-resistant Enterobacterales have become a global problem. We characterized extended-spectrum ß-lactamase (ESBL)-producing Enterobacterales from urinary tract infections cases from Allied Hospital Faisalabad, Pakistan. METHODS AND RESULTS: Eleven (22%, 11/50) ESBL-producing Enterobacterales (Escherichia coli; n = 10 and Enterobacter hormaechei; n = 1) were recovered and processed through VITEK-2, PCR, rep-PCR followed by whole-genome sequencing (WGS) of ESBL-producing Ent. hormaechei and carbapenem-resistant E. coli isolates. Plasmid transferability of blaNDM-1 -producers was assayed by conjugation experiments. All ESBL strains carried the blaCTX-M-15 gene. Of these blaCTX-M-15 producing E. coli, four also carried blaNDM-1 located on transferable plasmids. All E. coli strains belonged to ST448 and displayed similar genetic features including genes for antimicrobial resistance, heavy metal, biocides and virulence. Genomic features of a multidrug-resistant (MDR) Ent. hormaechei were also reported for the first time in Pakistan. CONCLUSION: Our findings indicate that blaNDM-1 producing E. coli ST448 is a multidrug, heavy metals and biocides-resistant strain. Therefore, the screening of these isolates may be effective in limiting the MDR bacteria spread in hospitalized patients and within the community. SIGNIFICANCE AND IMPACT OF THIS STUDY: Spread of multi-drug-resistant ESBL-producing bacteria in the clinical settings of Pakistan is a serious challenge and further limiting treatment options in the country. WGS could be used as a tool in the nationwide antibiotic surveillance programme to explore insights of spread and outbreak.

Eruca sativa seed napin structural insights and thorough functional characterization.

A potent napin protein has been thoroughly characterized from seeds of rocket salad (Eruca sativa). Eruca sativa napin (EsNap) was purified by ammonium sulfate precipitation (70%) and size-exclusion chromatography. Single intact 16 kDa EsNap band was reduced to 11 and 5 kDa bands respectively on SDS-PAGE. Nano LC-MS/MS yielded two fragments comprising of 26 residues which showed 100% sequence identity with napin-3 of Brassica napus. CD spectroscopy indicated a dominant α-helical structure of EsNap. Monodispersity of EsNap was verified by dynamic light scattering, which also confirmed the monomeric status with a corresponding hydrodynamic radius of 2.4 ± 0.2 nm. An elongated ab initio shape of EsNap was calculated based on SAXS data, with an Rg of 1.96 ± 0.1 nm. The ab initio model calculated by DAMMIF with P1 symmetry and a volume of approx. 31,100 nm3, which corresponded to a molecular weight of approximately 15.5 kDa. The comparison of the SAXS and ab initio modeling showed a minimized χ2-value of 1.87, confirming a similar molecular structure. A homology model was predicted using the coordinate information of Brassica napus rproBnIb (PDB ID: 1SM7). EsNap exhibited strong antifungal activity by significantly inhibiting the growth of Fusarium graminearum. EsNap also showed cytotoxicity against the hepatic cell line Huh7 and the obtained IC50 value was 20.49 µM. Further, strong entomotoxic activity was experienced against different life stages of stored grain insect pest T. castaneum. The result of this study shows insights that can be used in developing potential antifungal, anti-cancerous and insect resistance agents in the future using EsNap from E. sativa.

Rapid Identification of Common Secondary Metabolites of Medicinal Herbs Using High-Performance Liquid Chromatography with Evaporative Light Scattering Detector in Extracts.

The discovery and identification of novel natural products of medicinal importance in the herbal medicine industry becomes a challenge. The complexity of this process can be reduced by dereplication strategies. The current study includes a method based on high-performance liquid chromatography (HPLC), using the evaporative light scattering detector (ELSD) to identify the 12 most common secondary metabolites in plant extracts. Twelve compounds including rutin, taxifolin, quercetin, apigenin, kaempferol, betulinic acid, oleanolic acid, betulin, lupeol, stigmasterol, and ß-sitosterol were analyzed simultaneously. The polarity of the compounds varied greatly from highly polar (flavonoids) to non-polar (triterpenes and sterols). This method was also tested for HPLC-DAD and HPLC-ESI-MS/MS analysis. Oleanolic acid and ursolic acid could not be separated in HPLC-ELSD analysis but were differentiated using LC-ESI-MS/MS analysis due to different fragment ions. The regression values (R2 > 0.996) showed good linearity in the range of 50-1000 µg/mL for all compounds. The range of LOD and LOQ values were 7.76-38.30 µg/mL and 23.52-116.06 µg/mL, respectively. %RSD and % trueness values of inter and intraday studies were mostly <10%. This method was applied on 10 species of medicinal plants. The dereplication strategy has the potential to facilitate and shorten the identification process of common secondary metabolites in complex plant extracts.

Genomic features of a high-risk mcr-1.1-positive Escherichia coli ST10 isolated from cattle farm environment.

The environment plays an important role in the dissemination of clinically relevant antimicrobial-resistant bacteria and genes. In this study, we described genomic features of a plasmid-mediated colistin-resistant mcr-1-positive Escherichia coli strains (PK-3225) isolated from a dairy farm wastewater sample. After initial isolation and PCR detection of mcr-1-positive E. coli, whole-genome sequencing was performed using Illumina Hiseq 2500 followed by in silico analysis. Genetic context surrounding the mcr-1 gene was determined and SNP-based phylogenomic analysis was performed. Furthermore, plasmid analysis and conjugation assays were performed to determine transferability of mcr-1. E. coli PK-3225 belonged to ST10 and carried a broad resistome that included colistin (mcr-1), beta-lactam (blaTEM-IB), tetracycline (tetB), phenicol (catA1), macrolide (mdfA), trimethoprim (dfrA17), aminoglycosides (aadA5, aph(3")-Ib, aph(6)-Id), and sulphonamide (sul2) resistance genes. The draft genome of E. coli calculated as 4.9 Mbp. Conjugation experiment showed successful transfer of the mcr-1 gene to E. coli recipient strain J53. In silico analysis showed that mcr-1 was located on IncI2 plasmid of > 59 kb in length, with the nikB-mcr-1-pap2 gene array, and lack ISApl1. The phylogenomic analysis revealed that the PK-3225 was closely related to human ST10 E. coli from Brazil and USA. To our knowledge, this is the first draft genome sequence of mcr-1 carrying E. coli isolated from the farm environment in Pakistan. Considering the high burden of colistin resistance in Pakistan, presence of pandemic high-risk E. coli clones in the environment requires strict surveillance.

Genome and Secretome Analysis of Staphylotrichum longicolleum DSM105789 Cultured on Agro-Residual and Chitinous Biomass.

Staphylotrichum longicolleum FW57 (DSM105789) is a prolific chitinolytic fungus isolated from wood, with a chitinase activity of 0.11 ± 0.01 U/mg. We selected this strain for genome sequencing and annotation, and compiled its growth characteristics on four different chitinous substrates as well as two agro-industrial waste products. We found that the enzymatic mixture secreted by FW57 was not only able to digest pre-treated sugarcane bagasse, but also untreated sugarcane bagasse and maize leaves. The efficiency was comparable to a commercial enzymatic cocktail, highlighting the potential of the S. longicolleum enzyme mixture as an alternative pretreatment method. To further characterize the enzymes, which efficiently digested polymers such as cellulose, hemicellulose, pectin, starch, and lignin, we performed in-depth mass spectrometry-based secretome analysis using tryptic peptides from in-gel and in-solution digestions. Depending on the growth conditions, we were able to detect from 442 to 1092 proteins, which were annotated to identify from 134 to 224 putative carbohydrate-active enzymes (CAZymes) in five different families: glycoside hydrolases, auxiliary activities, carbohydrate esterases, polysaccharide lyases, glycosyl transferases, and proteins containing a carbohydrate-binding module, as well as combinations thereof. The FW57 enzyme mixture could be used to replace commercial enzyme cocktails for the digestion of agro-residual substrates.

High-Throughput Detection of an Alkaloidal Plant Metabolome in Plant Extracts Using LC-ESI-QTOF-MS.

Plant alkaloids represent a diverse group of nitrogen-containing natural products. These compounds are considered valuable in drug discovery and development. High-throughput identification of such plant secondary metabolites in complex plant extracts is essential for drug discovery, lead optimization, and understanding the biological pathway. The present study aims to rapidly identify different classes of alkaloids in plant extracts through the liquid chromatography with electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) approach using 161 isolated and purified alkaloids. These are biologically important unique alkaloids belonging to different sub-classes such as isoquinoline, quinoline, indole, tropane, pyridine, piperidine, quinolizidine, aporphine, steroidal, and terpenoid. The majority of these are not available commercially and are known to manifest valuable biological activities. Four pools of a maximum of 50 phytostandards each were prepared, based on their log P value to minimize co-elution for rapid and cost-effective analyses. MS/MS spectra were acquired in the positive ionization mode by using their [M + H]+ and/or [M + Na]+ with both the average collisional energy (25.5-62 eV) and individual collisional energies (10, 20, 30, and 40 eV). Accurate mass, high-resolution mass spectrometry (HR-MS) data, MS/MS data, and retention times were curated for each compound. The developed LC-MS/MS method was successfully used to interrogate and fast dereplicate alkaloids in 13 medicinal plant extracts and a herbal formulation. A total of 56 alkaloids were identified based on the reference standard retention times (RTs), HR-MS spectra, and/or MS/MS spectra. The MS data have been submitted to the MetaboLights online database (MTBLS2914). The mass spectrometric and chromatographic data will be useful for the discovery of new congeners and the study of biological pathways of alkaloids in the plant kingdom.

Insights into the genome and secretome of Fusarium metavorans DSM105788 by cultivation on agro-residual biomass and synthetic nutrient sources.

BACKGROUND: The transition to a biobased economy involving the depolymerization and fermentation of renewable agro-industrial sources is a challenge that can only be met by achieving the efficient hydrolysis of biomass to monosaccharides. In nature, lignocellulosic biomass is mainly decomposed by fungi. We recently identified six efficient cellulose degraders by screening fungi from Vietnam. RESULTS: We characterized a high-performance cellulase-producing strain, with an activity of 0.06 U/mg, which was identified as a member of the Fusarium solani species complex linkage 6 (Fusarium metavorans), isolated from mangrove wood (FW16.1, deposited as DSM105788). The genome, representing nine potential chromosomes, was sequenced using PacBio and Illumina technology. In-depth secretome analysis using six different synthetic and artificial cellulose substrates and two agro-industrial waste products identified 500 proteins, including 135 enzymes assigned to five different carbohydrate-active enzyme (CAZyme) classes. The F. metavorans enzyme cocktail was tested for saccharification activity on pre-treated sugarcane bagasse, as well as untreated sugarcane bagasse and maize leaves, where it was complemented with the commercial enzyme mixture Accellerase 1500. In the untreated sugarcane bagasse and maize leaves, initial cell wall degradation was observed in the presence of at least 196 µg/mL of the in-house cocktail. Increasing the dose to 336 µg/mL facilitated the saccharification of untreated sugarcane biomass, but had no further effect on the pre-treated biomass. CONCLUSION: Our results show that F. metavorans DSM105788 is a promising alternative pre-treatment for the degradation of agro-industrial lignocellulosic materials. The enzyme cocktail promotes the debranching of biopolymers surrounding the cellulose fibers and releases reduced sugars without process disadvantages or loss of carbohydrates.

Evaluation of cytotoxicity of areca nut and its commercial products on normal human gingival fibroblast and oral squamous cell carcinoma cell lines.

Consumption of areca nut products is the most common cause of oral cancers, particularly in South Asian countries. This study evaluates the cytotoxic and necrotizing effects of areca nut and its formulations on normal human gingival fibroblasts (HGF-1) and oral squamous cell carcinoma (OSCC, CAL-27) cell lines. Identification of various carcinogens and adulterants using LC-HR-ESI-MS/MS analysis was performed in the extracts of areca nut and its products. Apart from alkaloids and flavonoids, a major adulterant, saccharin was found in all the samples of chalia (one of the most common chewing products of areca nut) in the ranges between 1.697-7.170 mg/g of the sample. Cytotoxic studies showed that most of the areca nut products were found cytotoxic to HGF-1 cells while being relatively non-cytotoxic against CAL-27 cells, rather they promote the growth of cancer cells. Our findings revealed that the components of areca nut and its products were injurious to HGF-1 cells and caused necrosis, which may attenuate HGF-1 protection toward oral epithelial cells. Moreover, the non-cytotoxic effect of these products on cancer cell lines suggests further predisposal of the habitual chewers for developing oral carcinomas. This study will give a better understanding of the hazardous effects of areca nut products.

Metaproteomics reveals the structural and functional diversity of Dermatocarpon miniatum (L.) W. Mann. Microbiota.

Metaproteomics is a strategy to understand the taxonomy, functionality and metabolic pathways of the microbial communities. The relationship among the symbiotic microbiota in the entire lichen thallus, Dermatocarpon miniatum, was evaluated using the metaproteomic approach. Proteomic profiling using one-dimensional SDS-PAGE followed by LC-MS/MS analysis resulted in a total of 138 identified proteins via Mascot search against UniRef100 and Swiss-Prot databases. In addition to the fungal and algal partners, D. miniatum proteome encompasses proteins from prokaryotes, which is a multifarious community mainly dominated by cyanobacteria and proteobacteria. While proteins assigned to fungus were the most abundant (55 %), followed by protists (16 %), bacterial (13 %), plant (11 %), and viral (1 %) origin, whereas 4 % remained undefined. Various proteins were assigned to the different lichen symbionts by using Gene Ontology (GO) terms, e.g. fungal proteins involved in the oxidation-reduction process, protein folding and glycolytic process, while protists and bacterial proteins were involved in photosynthetic electron transport in photosystem II (PS II), ATP synthesis coupled proton transport, and carbon fixation. The presence of bacterial communities extended the traditional concept of fungal-algal lichen symbiotic interaction.

Significance of Preoperative Prone Position Computed Tomography in Free Hand Subaxial Cervical Pedicular Screwing

: The subaxial cervical pedicle screwing technique shows powerful biomechanical properties for posterior cervical fusion. When applying a pedicle screw using the freehand technique, it is essential to analyse cervical computed tomography and plan the surgery accordingly. Normal cervical computed tomography is usually performed in the supine position, whereas during surgery, the patient lies in a prone position. This fact leads us to suppose that radiological evaluations may yield misleading results. Our study aimed to investigate whether there is any superiority between preoperative preparation on computed tomography performed in the prone position and that performed in the supine position. Methods : This study included 17 patients (132 pedicle screws) who were recently operated on with cervical vertebral computed tomography in the prone position and 17 patients (136 pedicle screws) who were operated on by conventional cervical vertebral computed tomography as the control group. The patients in both groups were compared in terms of age, gender, pathological diagnosis, screw malposition and complications. A screw malposition evaluation was made according to the Gertzbein-Robbins scale. Results : No statistically significant difference was observed between the two groups regarding age, gender and pathological diagnosis. The screw malposition rate (from 11.1% to 6.9%, p<0.05), mean malposition distance (from 2.18 mm to 1.86 mm, p <0.05), and complications statistically significantly decreased in the prone position computed tomography group. Conclusion : Preoperative surgical planning by performing cervical vertebral computed tomography in the prone position reduces screw malposition and complications. Our surgical success increased with a simple modification that can be applied by all clinicians without creating additional radiation exposure or additional costs.