Validation of ultrasonography for non-invasive assessment of diaphragm function in muscular dystrophy

Abstract

Duchenne muscular dystrophy (DMD) is a severe, degenerative muscle disease caused by dystrophin mutations. The mdx mouse is a widely used animal model of DMD. The mdx diaphragm muscle most closely recapitulates key features of DMD muscles, including progressive fibrosis and considerable force loss. Diaphragm function in mdx mice is commonly evaluated by specific force measurements ex vivo. While useful, this method only measures force from a small muscle sample at one time point. Therefore, accurate assessment of diaphragm function in vivo would provide an important advancement to study the time-course of functional decline and treatment benefits. Here, we evaluated an ultrasonography technique for measuring time-dependent changes of diaphragm function in mdx mice. Diaphragm movement amplitude values for mdx mice were considerably lower than WT, decreased from 8 to 18 months of age, and correlated strongly with ex vivo specific force. We then investigated the time-course of diaphragm amplitude changes following administration of an adeno-associated viral vector expressing Flag-micro-dystrophin (AAV-μDys) to young adult mdx mice. Diaphragm amplitude peaked 4 weeks after AAV-μDys administration, and was 26% greater than control mdx mice at this time. This value decreased slightly to 21% above mdx controls after 12 weeks of treatment. Importantly, diaphragm amplitude again correlated strongly with ex vivo specific force. Also, diaphragm amplitude and specific force negatively correlated with fibrosis levels in the muscle. Together, our results validate diaphragm ultrasonography as a reliable technique for assessing time-dependent changes in dystrophic diaphragm function in vivo, and for evaluating potential therapies for DMD.

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Selective accumulation of biotin in arterial chemoreceptors- requirement for carotid body exocytotic dopamine secretion

Key points

• Biotin, a vitamin whose main role is to be a coenzyme for carboxylases, accumulates at unusually large amounts within cells of the carotid body (CB).
• In biotin-deficient rats biotin rapidly disappears from the blood; however it remains at relatively high levels in CB glomus cells. The CB contains high levels of mRNA for SLC5a6, a biotin transporter, and SLC19a3, a thiamine transporter regulated by biotin.
• Animals with biotin deficiency exhibit pronounced metabolic lactic acidosis. Remarkably, glomus cells from these animals have normal electrical and neurochemical properties. However they show a marked decrease in the size of quantal dopaminergic secretory events.
• Inhibitors of the vesicular monoamine transporter 2 (VMAT2) mimic the effect of biotin deficiency. In biotin-deficient animals, VMAT2 protein expression decreases in parallel with biotin depletion in CB cells.
• These data suggest that dopamine transport and/or storage in small secretory granules in glomus cells depend on biotin.

Abstract

Biotin is a water-soluble vitamin required for the function of carboxylases as well as for the regulation of gene expression. Here, we report that biotin accumulates in unusually large amounts in cells of arterial chemoreceptors, carotid body (CB) and adrenal medulla (AM). We show in a biotin-deficient rat model that the vitamin rapidly disappears from the blood and other tissues (including the AM), while remaining at relatively high levels in the CB. We have also observed that, in comparison with other peripheral neural tissues, CB cells contain high levels of SLC5a6, a biotin transporter, and SLC19a3, a thiamine transporter regulated by biotin. Biotin-deficient rats show a syndrome characterized by marked weight loss, metabolic lactic acidosis, aciduria and accelerated breathing with normal responsiveness to hypoxia. Remarkably, CB cells from biotin-deficient animals have normal electrophysiological and neurochemical (ATP levels and catecholamine synthesis) properties; however, they exhibit a marked decrease in the size of quantal catecholaminergic secretory events, which is not seen in AM cells. A similar differential secretory dysfunction is observed in CB cells treated with tetrabenazine, a selective inhibitor of the vesicular monoamine transporter 2 (VMAT2). VMAT2 is highly expressed in glomus cells (in comparison with VMAT1) and in biotin-deficient animals VMAT2 protein expression decreases in parallel with the decrease of biotin accumulated in CB cells. These data suggest that biotin has an essential role in the homeostasis of dopaminergic transmission modulating the transport and/or storage of transmitters within small secretory granules in glomus cells.

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Assays for the Degradation of Misfolded Proteins in Cells

This report describes protocols for measuring degradation rates of misfolded proteins by either western blot or fluorescence-based assays. The methods can be applied to analysis of other misfolded proteins and for high throughput screening.

http://ift.tt/2bJxZKB

An Easy Method for Plant Polysome Profiling

This protocol describes an easy method to extract and fractionate transcripts from plant tissues on the basis of the number of bound ribosomes. It allows a global estimate of translation activity and the determination of the translational status of specific mRNAs.

http://ift.tt/2bJqfZ6

Transcriptome Analysis of HepG2 Cells Expressing ORF3 from Swine Hepatitis E Virus to Determine the Effects of ORF3 on Host Cells

Hepatitis E virus- (HEV-) mediated hepatitis has become a global public health problem. An important regulatory protein of HEV, ORF3, influences multiple signal pathways in host cells. In this study, to investigate the function of ORF3 from the swine form of HEV (SHEV), high-throughput RNA-Seq-based screening was performed to identify the differentially expressed genes in ORF3-expressing HepG2 cells. The results were validated with quantitative real-time PCR and gene ontology was employed to assign differentially expressed genes to functional categories. The results indicated that, in the established ORF3-expressing HepG2 cells, the mRNA levels of CLDN6, YLPM1, APOC3, NLRP1, SCARA3, FGA, FGG, FGB, and FREM1 were upregulated, whereas the mRNA levels of SLC2A3, DKK1, BPIFB2, and PTGR1 were downregulated. The deregulated expression of CLDN6 and FREM1 might contribute to changes in integral membrane protein and basement membrane protein expression, expression changes for NLRP1 might affect the apoptosis of HepG2 cells, and the altered expression of APOC3, SCARA3, and DKK1 may affect lipid metabolism in HepG2 cells. In conclusion, ORF3 plays a functional role in virus-cell interactions by affecting the expression of integral membrane protein and basement membrane proteins and by altering the process of apoptosis and lipid metabolism in host cells. These findings provide important insight into the pathogenic mechanism of HEV.

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The Relationship between Sensory Latency and Amplitude

Publication date: Available online 28 August 2016
Source:Journal of Electromyography and Kinesiology
Author(s): Elliot B. Bodofsky, Stephen J. Cohen, Rohini J. Kumar, Adam Schindelheim, John Gaughan
PurposeTo prove that the relationship between sensory latencies and amplitudes is useful in determining the severity of neuropathies. This is achieved by deriving a mathematical relationship between sensory distal latency and amplitude. Determine whether sensory amplitudes below predicted correlate with a worse pathology. Procedures: Patients seen for Nerve Conduction Studies by the Department of Physical Medicine and Rehabilitation at Cooper University Hospital between 12/1/12 and 12/31/14 were invited to participate in a prospective database. The median, ulnar and sural sensory latencies and amplitudes were analyzed with both linear and power regression. Patients with amplitudes above and below the regression curve were compared for latency, amplitude and velocity of other nerves. Carpal Tunnel Patients were analyzed to determine whether Median sensory amplitude below predicted correlated with more severe disease. Results: For the Median nerve, Power Regression Analysis showed a stronger correlation (R²=0.54) than linear regression (R²=0.34). Patients with Median sensory amplitude below the power correlation curve showed significantly longer ulnar sensory latency, and lower sensory amplitude than those above. Carpal Tunnel Syndrome patients with Median sensory amplitude well below predicted by the power relationship showed more advanced disease. For the ulnar and sural sensory nerve, the difference between power and linear regression was not significant. Conclusions: A power regression curve correlates sensory latency and amplitude better than linear regression. The latency amplitude relationship correlates with other parameters of nerve function and severity of Carpal Tunnel Syndrome. This implies that below predicted sensory amplitude may indicate worse disease, and could be a useful diagnostic tool.



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Isolation, Culture and Transduction of Adult Mouse Cardiomyocytes

This protocol describes a step-by-step method for the reproducible isolation and long-term culture of adult mouse cardiomyocytes with high yield, purity, and viability.

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Galactose-Deficient IgA1 as a Candidate Urinary Polypeptide Marker of IgA Nephropathy?

In patients with IgA nephropathy (IgAN), circulatory IgA1 and IgA1 in mesangial deposits contain elevated amounts of galactose-deficient IgA1 (Gd-IgA1). We hypothesized that a fraction of Gd-IgA1 from the glomerular deposits and/or circulation may be excreted into the urine and thus represent a disease-specific biomarker. Levels of urinary IgA and Gd-IgA1 were determined in 207 patients with IgAN, 205 patients with other renal diseases, and 57 healthy controls, recruited in USA, Japan, and Italy. Urinary IgA was similarly elevated in patients with IgAN and renal-disease controls compared with healthy controls. However, urinary Gd-IgA1 levels were higher in patients with IgAN (IgAN, ; disease controls, units/mg urinary creatinine; ). Lectin western blotting data confirmed these results. In IgAN patients, levels of urinary Gd-IgA1 correlated with proteinuria (). When we purified IgA from serum and urine of an IgAN patient, the relative proportion of Gd-IgA1 to total IgA1 was higher in the urine compared with serum, suggesting selective excretion of Gd-IgA1 in IgAN. In summary, urinary excretion of Gd-IgA1 was elevated in patients with IgAN and the urinary Gd-IgA1 levels correlated with proteinuria. Urinary Gd-IgA1 may thus represent a disease-specific biomarker of IgAN.

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Label-free quantitative proteomics reveals differentially expressed proteins in high risk childhood acute lymphoblastic leukemia

Publication date: 6 January 2017
Source:Journal of Proteomics, Volume 150
Author(s): Gang Xu, Zhijie Li, Lili Wang, Fang Chen, Zuofei Chi, Min Gu, Shuang li, Di Wu, Jianing Miao, Yi Zhang, Liangchun Hao, Yang Fan
Acute lymphoblastic leukemia (ALL) is the most common pediatric cancer. B-ALL is the most common type in pediatric ALL. Risk stratification is critical for setting on chemo-therapeutic regimen that has great impact on the survival rate. But the risk allocation schemas at present were not satisfied. We performed a proteomic study expecting to figure out the critical altered proteins which can indicate the risk rank. We depicted 86 differently expressed proteins in the high-risk childhood B-ALL, and 35 proteins were predicted to have directive interactions. We validated five identified proteins by immunoblot using specimens same as proteomics, and others different from that. We found the differently expressed proteins participated in pre-mRNA spicing, DNA damage response, and stress response which indicated different events happened in the high risk B-ALL. Our result provided new information for children B-ALL. It might aid more accurate risk stratification and might also be valuable to find new therapeutic targets.Biological significanceTo our knowledge, this is the first proteomic analysis comparing the differentially expressed proteins between high risk and low risk of childhood B-ALL by a label-free quantitative proteomics. We found in high risk B-ALL, the aberrant evens happened in pre-mRNA splicing, DNA damage response, and stress response. This study reveals new insights in the high risk B-ALL, and might also be valuable to identify the high-risk more accurately, as well as to find new therapeutic targets.

Graphical abstract

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Effervescent Granules Prepared Using Eucommia ulmoides Oliv. and Moso Bamboo Leaves: Hypoglycemic Activity in HepG2 Cells

Eucommia ulmoides Oliv. (E. ulmoides Oliv.) and moso bamboo (Phyllostachys pubescens) leaves are used as folk medicines in central-western China to treat diabetes. To investigate the hypoglycemic activity of the effervescent granules prepared using E. ulmoides Oliv. and moso bamboo leaves (EBEG) in HepG2 cells, EBEG were prepared with 5% of each of polysaccharides and chlorogenic acids from moso bamboo and E. ulmoides Oliv. leaves, respectively. HepG2 cells cultured in a high-glucose medium were classified into different groups. The results displayed EBEG-treated cells showed better glucose utilization than the negative controls; thus, the hypoglycemic effect of EBEG was much greater than that of granules prepared using either component alone, thereby indicating that this effect was due to a synergistic action of the components. Further, glucose consumption levels in the cells treated with EBEG (156.35% at 200 μg/mL) and the positive controls (metformin, 162.29%; insulin, 161.52%) were similar. Thus, EBEG exhibited good potential for use as a natural antidiabetic agent. The hypoglycemic effect of EBEG could be due to the synergistic action of polysaccharides from the moso bamboo leaves and chlorogenic acids from E. ulmoides Oliv. leaves via the inhibition of alpha-glucosidase and glucose-6-phosphate displacement enzyme.

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Preservation of Cognitive Function by Lepidium meyenii (Maca) Is Associated with Improvement of Mitochondrial Activity and Upregulation of Autophagy-Related Proteins in Middle-Aged Mouse Cortex

Maca has been used as a foodstuff and a traditional medicine in the Andean region for over 2,000 years. Recently the neuroprotective effects of maca also arouse interest of researchers. Decrease in mitochondrial function and decline in autophagy signaling may participate in the process of age-related cognitive decline. This study aimed to investigate if maca could improve cognitive function of middle-aged mice and if this effect was associated with improvement of mitochondrial activity and modulation of autophagy signaling in mouse cortex. Fourteen-month-old male ICR mice received maca powder administered by gavage for five weeks. Maca improved cognitive function, motor coordination, and endurance capacity in middle-aged mice, accompanied by increased mitochondrial respiratory function and upregulation of autophagy-related proteins in cortex. Our findings suggest that maca is a newly defined nutritional plant which can improve mitochondrial function and upregulate autophagy-related proteins and may be an effective functional food for slowing down age-related cognitive decline.

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Curative Effects of Suhuang Zhike Capsule on Postinfectious Cough: A Meta-Analysis of Randomized Trials

Objective. In this paper, we intended to systematically evaluate the efficacy of Suhuang Zhike Capsule (SZC) on postinfectious cough (PIC) in adults (age > 18). Methods. MEDLINE (PubMed), Chinese National Knowledge Infrastructure (CNKI), Cqvip Database (VIP), and Wanfang Database were researched for the randomized controlled trials (RCTs) of SZC for PIC. The search was limited to human studies, using the search keywords or free-text terms "cough," "post-infectious cough," "postinfectious cough," "post-cold cough," "postviral cough," "postcold cough," "Suhuang Zhike capsule," "Chinese Medicine," and "randomized clinical trials". Two reviewers individually extracted data from the included RCTs and then the extracted data were analyzed using Review Manager 5.3 software. Results. Seven RCTs involving 573 patients entered the inclusion criteria. Findings suggested that, compared with western conventional medicine (WCM) and other Chinese medicine, SZC could effectively improve the efficacy rate (OR 2.68, 95%  CI, 1.48–4.84, ; OR 4.86, 95%  CI, 1.50–15.73, , separately). Moreover, SZC could also improve the efficacy rate of Chinese medicine symptom (MD −0.74, 95%  CI, −1.46~−0.02, ). However, in terms of cough relief time, more evidence is needed to prove that SZC have an earlier antitussive effect (MD −1.31, 95%  CI, −3.06~0.45, ). Conclusion. The current evidence shows that SZC is effective in the treatment of PIC in adults and can significantly improve the effective rate of Chinese medicine symptoms.

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Coronary Artery Calcification Is Related to Inflammation in Rheumatoid Arthritis: A Long-Term Follow-Up Study

Objective. A long-term follow-up of patients with rheumatoid arthritis (RA) to evaluate factors related to coronary artery calcification (CAC). Methods. All 22 eligible patients (4 males/18 females, mean age 65 years, and RA-duration 30–36 years) from the original (baseline; ) study of atherosclerosis were included. Inflammation, cardiovascular risk factors, and biomarkers were measured at baseline. At follow-up 13 years later, CAC was assessed by computed tomography (CT) and the grade of inflammation was measured. Multivariate analysis of differences between patients with low (0–10) and high CAC (>10) was done by orthogonal projection to latent structures (OPLS). Results. Ten patients had CAC 0–10 and 12 had >10 (range 18–1700). Patients with high CAC had significantly higher ESR (24.3 versus 9.9 mm/h) and swollen joint count (2 versus 0). The OPLS models discriminated between patients having high or low CAC. With only baseline variables, the sensitivity was 73% and the specificity 82%. The model that also included inflammatory variables from follow-up had a sensitivity of 89% and a specificity of 85%. Exclusion of baseline intima media thickness and plaque from the latter model modestly reduced the accuracy (sensitivity 80% and specificity 83%). Conclusions. CAC is related to inflammation in patients with RA.

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Embryoid Body-Explant Outgrowth Cultivation from Induced Pluripotent Stem Cells in an Automated Closed Platform

Automation of cell culture would facilitate stable cell expansion with consistent quality. In the present study, feasibility of an automated closed-cell culture system "P 4C S" for an embryoid body- (EB-) explant outgrowth culture was investigated as a model case for explant culture. After placing the induced pluripotent stem cell- (iPSC-) derived EBs into the system, the EBs successfully adhered to the culture surface and the cell outgrowth was clearly observed surrounding the adherent EBs. After confirming the outgrowth, we carried out subculture manipulation, in which the detached cells were simply dispersed by shaking the culture flask, leading to uniform cell distribution. This enabled continuous stable cell expansion, resulting in a cell yield of 3.1 × 107. There was no evidence of bacterial contamination throughout the cell culture experiments. We herewith developed the automated cultivation platform for EB-explant outgrowth cells.

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Genus Tinospora: Ethnopharmacology, Phytochemistry, and Pharmacology

The genus Tinospora includes 34 species, in which several herbs were used as traditional medicines by indigenous groups throughout the tropical and subtropical parts of Asia, Africa, and Australia. The extensive literature survey revealed Tinospora species to be a group of important medicinal plants used for the ethnomedical treatment of colds, headaches, pharyngitis, fever, diarrhea, oral ulcer, diabetes, digestive disorder, and rheumatoid arthritis. Indian ethnopharmacological data points to the therapeutic potential of the T. cordifolia for the treatment of diabetic conditions. While Tinospora species are confusing in individual ingredients and their mechanisms of action, the ethnopharmacological history of those plants indicated that they exhibit antidiabetic, antioxidation, antitumor, anti-inflammation, antimicrobial, antiosteoporosis, and immunostimulation activities. While the clinical applications in modern medicine are lacking convincing evidence and support, this review is aimed at summarizing the current knowledge of the traditional uses, phytochemistry, biological activities, and toxicities of the genus Tinospora to reveal its therapeutic potentials and gaps, offering opportunities for future researches.

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Frailty and influenza vaccine effectiveness

Publication date: 7 September 2016
Source:Vaccine, Volume 34, Issue 39
Author(s): Sheena G. Sullivan, Benjamin J. Cowling, Sander Greenland




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Reply to Sullivan et al.

Publication date: 7 September 2016
Source:Vaccine, Volume 34, Issue 39
Author(s): Qingxia Chen, Hui Nian, Yuwei Zhu, H. Keipp Talbot




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Editorial Board/Aims and Scope

Publication date: 7 September 2016
Source:Vaccine, Volume 34, Issue 39





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Corrigendum to “Use of the nonavalent HPV vaccine in individuals previously fully or partially vaccinated with bivalent or quadrivalent HPV vaccines” [Vaccine 34 (2016) 757–761]

Publication date: 7 September 2016
Source:Vaccine, Volume 34, Issue 39
Author(s): Pierre Van Damme, Paolo Bonanni, F. Xavier Bosch, Elmar Joura, Susanne Krüger Kjaer, Chris J.L.M. Meijer, Karl-Ulrich Petry, Benoit Soubeyrand, Thomas Verstraeten, Margaret Stanley




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Diazoxide Attenuates Postresuscitation Brain Injury in a Rat Model of Asphyxial Cardiac Arrest by Opening Mitochondrial ATP-Sensitive Potassium Channels

Objective. We investigated whether and how diazoxide can attenuate brain injury after cardiopulmonary resuscitation (CPR) by selective opening of mitochondrial ATP-sensitive potassium (mitoKATP) channels. Methods. Adult male Sprague-Dawley rats with induced cerebral ischemia ( per group) received an intraperitoneal injection of 0.1% dimethyl sulfoxide (1 mL; vehicle group), diazoxide (10 mg/kg; DZ group), or diazoxide (10 mg/kg) plus 5-hydroxydecanoate (5 mg/kg; DZ + 5-HD group) 30 min after CPR. The control group (sham group, ) underwent sham operation, without cardiac arrest. Mitochondrial respiratory control rate (RCR) was determined. Brain cell apoptosis was assessed using TUNEL staining. Expression of Bcl-2, Bax, and protein kinase C epsilon (PKCε) in the cerebral cortex was determined by Western blotting and immunohistochemistry. Results. The neurological deficit scores (NDS) in the vehicle group decreased significantly at 24 h and 48 h after CPR. Diazoxide significantly improved NDS and mitochondrial RCR after CPR at both time points; 5-HD cotreatment abolished these effects. Diazoxide decreased TUNEL-positive cells following CPR, upregulated Bcl-2 and PKCε, downregulated Bax, and increased the Bcl-2/Bax ratio; 5-HD cotreatment reversed these effects. Conclusions. Diazoxide attenuates postresuscitation brain injury, protects mitochondrial function, inhibits brain cell apoptosis, and activates the PKC pathway by opening mitoKATP channels.

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ProFold: Protein Fold Classification with Additional Structural Features and a Novel Ensemble Classifier

Protein fold classification plays an important role in both protein functional analysis and drug design. The number of proteins in PDB is very large, but only a very small part is categorized and stored in the SCOPe database. Therefore, it is necessary to develop an efficient method for protein fold classification. In recent years, a variety of classification methods have been used in many protein fold classification studies. In this study, we propose a novel classification method called proFold. We import protein tertiary structure in the period of feature extraction and employ a novel ensemble strategy in the period of classifier training. Compared with existing similar ensemble classifiers using the same widely used dataset (DD-dataset), proFold achieves 76.2% overall accuracy. Another two commonly used datasets, EDD-dataset and TG-dataset, are also tested, of which the accuracies are 93.2% and 94.3%, higher than the existing methods. ProFold is available to the public as a web-server.

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Simultaneous bioimaging recognition of Al3+ and Cu2+ in living-cell, and further detection of F− and S2− by a simple fluorogenic benzimidazole-based chemosensor

Publication date: 1 December 2016
Source:Talanta, Volume 161
Author(s): Haiyang Liu, Bibo Zhang, Chunyan Tan, Feng Liu, Jiakun Cao, Ying Tan, Yuyang Jiang
A simple Schiff base (BMSA) prepared from salicylaldehyde and 2-(1H-benzo[d]imidazol-2-yl)aniline was evaluated as an efficient fluorescent chemosensor for the selective recognition of Al³⁺and Cu²⁺ over other common metal ions. This sensor could detect Al3⁺ in CH3OH/PBS with distinct emission red-shift (the detection limit 0.31μM)and Cu²⁺in CH3OH/Tris-HCL (the detection limit 0.54μM) with obvious fluorescence quenching. The obtained BMSA-Al³⁺ and BMSA-Cu²⁺ complexes could act as cascade sensors for detecting F⁻ and S²⁻, respectively. The recognizing behavior of BMSA toward Al³⁺and Cu²⁺ has been investigated in detail through Job's Plot, FT-IR NMR, and HRMS analysis. Moreover, this chemosensor was verified to be of low cytotoxicity and good imaging characteristics for the detection of Al³⁺ and Cu²⁺, and further for the recognition of F⁻ and S²⁻ in living cells, suggesting that BMSA was proved to be a useful tool for tracking Al3⁺/Cu²⁺and F⁻/S²⁻ ions in vivo.

Graphical abstract

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Neural networks applied to characterize blends containing refined and extra virgin olive oils

Publication date: 1 December 2016
Source:Talanta, Volume 161
Author(s): Regina Aroca-Santos, John C. Cancilla, Enrique S. Pariente, José S. Torrecilla
The identification and quantification of binary blends of refined olive oil with four different extra virgin olive oil (EVOO) varietals (Picual, Cornicabra, Hojiblanca and Arbequina) was carried out with a simple method based on combining visible spectroscopy and non-linear artificial neural networks (ANNs).The data obtained from the spectroscopic analysis was treated and prepared to be used as independent variables for a multilayer perceptron (MLP) model. The model was able to perfectly classify the EVOO varietal (100% identification rate), whereas the error for the quantification of EVOO in the mixtures containing between 0% and 20% of refined olive oil, in terms of the mean prediction error (MPE), was 2.14%. These results turn visible spectroscopy and MLP models into a trustworthy, user-friendly, low-cost technique which can be implemented on-line to characterize olive oil mixtures containing refined olive oil and EVOOs.

Graphical abstract

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Discrimination and quantification of two isomeric antineoplastic drugs by rapid and non-invasive analytical control using a handheld Raman spectrometer

Publication date: 1 December 2016
Source:Talanta, Volume 161
Author(s): L.M.M. Lê, A. Tfayli, J. Zhou, P. Prognon, A. Baillet-Guffroy, E. Caudron
Raman spectroscopy is a rapid, non-destructive and non-invasive method that is a promising tool for real-time analytical control of drug concentrations. This study evaluated a handheld Raman device to discriminate and quantify two isomeric drugs used to treat cancer. Doxorubicin (DOXO) and epirubicin (EPIR) samples were analyzed at therapeutic concentrations from 0.1 to 2mg/mL (n=90) and 0.08–2mg/mL (n=90) by non-invasive measurements using a portable Raman spectrometer.The discrimination of these two molecules was demonstrated for all concentrations (n=180) by qualitative analysis using partial least square discriminant analysis (PLS-DA) with 100% classification accuracy, sensitivity and specificity and 0% error rate. For each molecule, quantitative analyses were performed using PLS regression. The validity of the model was evaluated using root mean square error of cross validation (RMSECV) and prediction (RMSEP) that furnished 0.05 and 0.02mg/mL for DOXO and 0.17 and 0.16mg/mL for EPIR after pretreatment optimization. Based on the accuracy profile, the linearity range was from 1.256 to 2.000mg/mL for DOXO (R²=0.9988) and from 0.553 to 2.000mg/Ml for EPIR (R²=0.9240) and repeatability (CV% max of 1.8% for DOXO and 3.2% for EPIR) and intermediate precision (CV% max of 2.8% for DOXO and 4.5% for EPIR) were both acceptable.Despite the narrow validated concentration range for quantitative analysis, this study shows the potential of a handheld Raman spectrometer coupled to chemometric approaches for real-time quantification of cytotoxic drugs, as well for discriminating between two drugs with similar UV absorption profiles. Finally, the use of a handheld spectrometer with the possibility of a direct measurement of substances in containers is a potentially valuable tool for combining patient safety with security of healthcare workers.

Graphical abstract

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