scholarly journals An Electrochemical Chip to Monitor In Vitro Glycation of Proteins and Screening of Antiglycation Potential of Drugs

Pharmaceutics ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 1011
Author(s):  
Zeeshan A. Khan ◽  
Seungkyung Park
Keyword(s):  
Inhibitory Concentration ◽  
Strong Relationship ◽  
Protein Glycation ◽  
Capacitance Measurement ◽  
Efficacy Evaluation ◽  
Chronic Complications ◽  
Glycation End Products ◽  
Significant Change ◽  
Selection Of

Hyperglycemia and the production of advanced glycation end products (AGEs) are the primary factors for the development of chronic complications in diabetes. The level of protein glycation is proportional to the glucose concentration and represents mean glycemia. In this study, we present an electrochemical chip-based method for in vitro glycation monitoring and the efficacy evaluation of an antiglycation compound. An electrochemical chip consisting of five microchambers and embedded microelectrodes was designed for parallel measurements of capacitance signals from multiple solutions at different concentrations. The feasibility of glycation monitoring was then investigated by measuring the capacitance signal at 0.13 MHz with bovine serum albumin and gelatin samples in the presence of various glucose concentrations over 28 days. A significant change in the capacitance due to protein glycation was observed through measurements conducted within 30 s and 21 days of incubation. Finally, we demonstrated that the chip-based capacitance measurement can be utilized for the selection of an antiglycation compound by supplementing the protein solution and hyperglycemic concentration of glucose with an inhibitory concentration of the standard antiglycation agent aspirin. The lack of a significant change in the capacitance over 28 days proved that aspirin is capable of inhibiting protein glycation. Thus, a strong relationship exists between glycation and capacitance, suggesting the application of an electrochemical chip for evaluating glycation and novel antiglycation agents.

scholarly journals Protein Glycation in Diabetes as Determined by Mass Spectrometry

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Annunziata Lapolla ◽  
Laura Molin ◽  
Pietro Traldi
Keyword(s):  
Mass Spectrometry ◽  
Degradation Products ◽  
Glycated Haemoglobin ◽  
Maldi Mass Spectrometry ◽  
Enzymatic Digestion ◽  
Protein Glycation ◽  
Chronic Complications ◽  
Glycation End Products

Diabetes is a common endocrine disorder characterized by hyperglycemia leading to nonenzymatic glycation of proteins, responsible for chronic complications. The development of mass spectrometric techniques able to give highly specific and reliable results in proteome field is of wide interest for physicians, giving them new tools to monitor the disease progression and the possible complications related to diabetes, as well as the effectiveness of therapeutic treatments. This paper reports and discusses some of the data pertaining protein glycation in diabetic subjects obtained by matrix-assisted laser desorption ionization (MALDI) mass spectrometry (MS). The preliminary studies carried out byin vitroprotein glycation experiments show clear differences in molecular weight of glycated and unglycated proteins. Then, the attention was focused on plasma proteins human serum albumin (HSA) and immunoglobulin G (IgG). Enzymatic degradation products ofin vitroglycated HSA were studied in order to simulate thein vivoenzymatic digestion of glycated species by the immunological system leading to the highly reactive advanced glycation end-products (AGEs) peptides. Further studies led to the evaluation of glycated Apo A-I and glycated haemoglobin levels. A different MALDI approach was employed for the identification of markers of disease in urine samples of healthy, diabetic, nephropathic, and diabetic-nephropathic subjects.

scholarly journals The Inhibitory Effect ofPrunella vulgarisL. on Aldose Reductase and Protein Glycation

2012 ◽  
Vol 2012 ◽  
pp. 1-7 ◽  
Author(s):  
Hong Mei Li ◽  
Jin Kyu Kim ◽  
Jai Man Jang ◽  
Sang Oh Kwon ◽  
Cheng Bi Cui ◽  
...  
Keyword(s):  
Aldose Reductase ◽  
Inhibitory Effect ◽  
Protein Glycation ◽  
Prunella Vulgaris ◽  
Inhibitory Effects ◽  
Advanced Glycation ◽  
High Concentration ◽  
Potent Inhibition ◽  
Glycation End Products

To evaluate the aldose reductase (AR) enzyme inhibitory ability ofPrunella vulgarisL. extract, six compounds were isolated and tested for their effects. The components were subjected toin vitrobioassays to investigate their inhibitory assays using rat lens aldose reductase (rAR) and human recombinant AR (rhAR). Among them, caffeic acid ethylene ester showed the potent inhibition, with the IC50values of rAR and rhAR at3.2±0.55 μM and12.58±0.32 μM, respectively. In the kinetic analyses using Lineweaver-Burk plots of 1/velocity and 1/concentration of substrate, this compound showed noncompetitive inhibition against rhAR. Furthermore, it inhibited galactitol formation in a rat lens incubated with a high concentration of galactose. Also it has antioxidative as well as advanced glycation end products (AGEs) inhibitory effects. As a result, this compound could be offered as a leading compound for further study as a new natural products drug for diabetic complications.

In Vitro Analytical System for Determining the Ability of Antibiotics at Residue Levels to Select for Resistance in Bacteria

1988 ◽  
Vol 71 (2) ◽  
pp. 295-298
Author(s):  
Marietta Suebrady ◽  
Robert J Strobel ◽  
Stanley E Katz
Keyword(s):  
Analytical Procedure ◽  
Inhibitory Concentration ◽  
Enteric Bacteria ◽  
E Coli ◽  
Resistant Population ◽  
Test Organisms ◽  
Residue Levels ◽  
Analytical System ◽  
Selection Of

Abstract An analytical procedure, based on the concept that exposure of bacteria to antibiotics will result in the selection of a resistant population, was developed. Two strains of enteric bacteria, Escherichia coli CS-1 and Enterobacter cloacae B520, which are sensitive to a wide variety of antibiotics, were used as the test organisms. E. coli CS-1 were exposed to 1.00 μg antibiotic or antimicrobial/mL; E. cloacae B520 were exposed to 0.01, 0.10, 0.50,1.00, and 5.00 μg/mL. Both organisms developed increased resistance to other antibiotics after exposure to chlortetracycline and oxytetracycline, as measured by the minimum inhibitory concentration (MIC). E. cloacae B520 showed increased resistance to ampicillin, oxytetracycline, and chloramphenicol after exposure to levels as low as 0.10 μg/mL. Exposure to streptomycin, sulfamethazine, tylosin, bacitracin, flavomycin, virginiamycin, and monensin at levels of 1.00 μg/mL did not increase the MIC. Exposure to 5.00 *tg streptomycin, sulfamethazine, tylosin, and monensin/mL increased the MIC ofE. cloacae to one of the antibiotic markers. These increased MICs exceeded the 95% confidence limits of the MIC values of the unexposed organisms.

scholarly journals Rezafungin—Mechanisms of Action, Susceptibility and Resistance: Similarities and Differences with the Other Echinocandins

2020 ◽  
Vol 6 (4) ◽  
pp. 262
Author(s):  
Guillermo Garcia-Effron
Keyword(s):  
Inhibitory Concentration ◽  
Similar Frequency ◽  
Resistant Strains ◽  
Long Acting ◽  
Synthase Inhibitor ◽  
New Generation ◽  
Susceptibility And Resistance ◽  
In Vitro Data ◽  
Selection Of

Rezafungin (formerly CD101) is a new β-glucan synthase inhibitor that is chemically related with anidulafungin. It is considered the first molecule of the new generation of long-acting echinocandins. It has several advantages over the already approved by the Food and Drug Administration (FDA) echinocandins as it has better tissue penetration, better pharmacokinetic/phamacodynamic (PK/PD) pharmacometrics, and a good safety profile. It is much more stable in solution than the older echinocandins, making it more flexible in terms of dosing, storage, and manufacturing. These properties would allow rezafungin to be administered once-weekly (intravenous) and to be potentially administered topically and subcutaneously. In addition, higher dose regimens were tested with no evidence of toxic effect. This will eventually prevent (or reduce) the selection of resistant strains. Rezafungin also has several similarities with older echinocandins as they share the same in vitro behavior (very similar Minimum Inhibitory Concentration required to inhibit the growth of 50% of the isolates (MIC50) and half enzyme maximal inhibitory concentration 50% (IC50)) and spectrum, the same target, and the same mechanisms of resistance. The selection of FKS mutants occurred at similar frequency for rezafungin than for anidulafungin and caspofungin. In this review, rezafungin mechanism of action, target, mechanism of resistance, and in vitro data are described in a comparative manner with the already approved echinocandins.

In vitro inhibition of protein glycation and advanced glycation end products formation by hydroethanolic extract and two fractions of Simaba trichilioides roots

2018 ◽  
Vol 34 (16) ◽  
pp. 2389-2393
Author(s):  
Bruno Pereira Motta ◽  
Anderson Kiyoshi Kaga ◽  
Juliana Oriel Oliveira ◽  
Maiara Destro Inacio ◽  
Cledson Ferreira da Silva ◽  
...  
Keyword(s):  
Advanced Glycation End Products ◽  
Protein Glycation ◽  
Advanced Glycation ◽  
Glycation End Products ◽  
End Products ◽  
In Vitro Inhibition ◽  
Hydroethanolic Extract

scholarly journals An axenic amastigote system for drug screening.

1997 ◽  
Vol 41 (4) ◽  
pp. 818-822 ◽  
Author(s):  
H L Callahan ◽  
A C Portal ◽  
R Devereaux ◽  
M Grogl
Keyword(s):  
Drug Screening ◽  
Inhibitory Concentration ◽  
Rapid Testing ◽  
Intracellular Amastigotes ◽  
Axenic Amastigotes ◽  
Parasite Stage ◽  
Intracellular Amastigote ◽  
Selection Of

Currently available primary screens for selection of candidate antileishmanial compounds are not ideal. The choices include screens that are designed to closely reflect the situation in vivo but are labor-intensive and expensive (intracellular amastigotes and animal models) and screens that are designed to facilitate rapid testing of a large number of drugs but do not use the clinically relevant parasite stage (promastigote model). The advent of successful in vitro culture of axenic amastigotes permits the development of a primary screen which is quick and easy like the promastigote screen but still representative of the situation in vivo, since it uses the relevant parasite stage. We have established an axenic amastigote drug screening system using a Leishmania mexicana strain (strain M379). A comparison of the 50% inhibitory concentration (IC50) drug sensitivity profiles of M379 promastigotes, intracellular amastigotes, and axenic amastigotes for six clinically relevant antileishmanial drugs (sodium stibogluconate, meglumine antimoniate, pentamidine, paromomycin, amphotericin B, WR6026) showed that M379 axenic amastigotes are a good model for a primary drug screen. Promastigote and intracellular amastigote IC50s differed for four of the six drugs tested by threefold or more; axenic amastigote and intracellular amastigote IC50s differed by twofold for only one drug. This shows that the axenic amastigote susceptibility to clinically used reference drugs is comparable to the susceptibility of amastigotes in macrophages. These data also suggest that for the compounds tested, susceptibility is intrinsic to the parasite stage. This contradicts previous hypotheses that suggested that the activities of antimonial agents against intracellular amastigotes were solely a function of the macrophage.

scholarly journals Improved Methods for the Rapid Formation and Prevention of Advanced Glycation End Products (AGEs) In Vitro by Coupling to the Hypoxanthine/Xanthine Oxidase Assay System

Biomedicines ◽  
2018 ◽  
Vol 6 (3) ◽  
pp. 88 ◽  
Author(s):  
Samuel Marques ◽  
Teresa Trevisan ◽  
Carlos Maia ◽  
Andrea Breuer ◽  
Robert Owen
Keyword(s):  
Xanthine Oxidase ◽  
Advanced Glycation End Products ◽  
High Capacity ◽  
Protein Glycation ◽  
Advanced Glycation ◽  
Leaf Extracts ◽  
Reactive Carbonyl Species ◽  
Glycation End Products ◽  
End Products

Advanced glycation end products (AGEs) represent a set of molecules that contribute directly to the initiation and aggravation of diseases associated with ageing. AGEs are produced by the reaction between reducing sugars (or α-dicarbonyl compounds), proteins, and amino acid residues. Previous in vitro methods using non-enzymatic procedures described in the literature require an incubation period of 1–3 weeks to generate AGEs. In this study, the reaction time for the formation of AGEs (48 and 3 h) was significantly reduced by adaptation of methods previously described in the literature and coupling them to the free radical generation system termed hypoxanthine/xanthine oxidase assay. The incorporation of this assay into the experimental system accelerated the production of AGEs as a result of the formation of reactive oxygen species (ROS), as shown by increased fluorescence. The capacity of different classes of chemical compounds (aminoguanidine, chlorogenic acid, rutin, and methanol extracts of Hancornia speciosa Gomes) to inhibit protein glycation by acting as scavenging agents of α-dicarbonyl species was evaluated. Aminoguanidine and, especially, rutin identified in the leaf extracts of H. speciosa Gomes showed a high capacity to act as scavengers of reactive carbonyl species RCS-trapping, resulting in the inhibition of AGEs formation.

scholarly journals Antibody-like proteins that capture and neutralize SARS-CoV-2

2020 ◽  
Vol 6 (42) ◽  
pp. eabd3916 ◽  
Author(s):  
T. Kondo ◽  
Y. Iwatani ◽  
K. Matsuoka ◽  
T. Fujino ◽  
S. Umemoto ◽  
...  
Keyword(s):  
High Speed ◽  
In Vitro Selection ◽  
Nasal Swab ◽  
Inhibitory Concentration ◽  
Rapid Development ◽  
Spike Protein ◽  
Emerging Pathogens ◽  
High Affinity ◽  
Selection Of

To combat severe acute respiratory syndrome–related coronavirus 2 (SARS-CoV-2) and any unknown emerging pathogens in the future, the development of a rapid and effective method to generate high-affinity antibodies or antibody-like proteins is of critical importance. We here report high-speed in vitro selection of multiple high-affinity antibody-like proteins against various targets including the SARS-CoV-2 spike protein. The sequences of monobodies against the SARS-CoV-2 spike protein were successfully procured within only 4 days. Furthermore, the obtained monobody efficiently captured SARS-CoV-2 particles from the nasal swab samples of patients and exhibited a high neutralizing activity against SARS-CoV-2 infection (half-maximal inhibitory concentration, 0.5 nanomolar). High-speed in vitro selection of antibody-like proteins is a promising method for rapid development of a detection method for, and of a neutralizing protein against, a virus responsible for an ongoing, and possibly a future, pandemic.

scholarly journals Influence of Piracetam on Gliclazide—Glycated Human Serum Albumin Interaction. A Spectrofluorometric Study

Molecules ◽  
2018 ◽  
Vol 24 (1) ◽  
pp. 111 ◽  
Author(s):  
Agnieszka Szkudlarek ◽  
Jadwiga Pożycka ◽  
Małgorzata Maciążek-Jurczyk
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Glycated Albumin ◽  
Human Albumin ◽  
Protein Glycation ◽  
Monitoring Therapy ◽  
Glycation End Products ◽  
Local Accumulation

Advanced Glycation End-Products (AGEs) are created in the last step of protein glycation and can be a factor in aging and in the development or worsening of many degenerative diseases (diabetes, chronic kidney disease, atherosclerosis, Alzheimer’s disease, etc.). Albumin is the most susceptible to glycation plasma protein. Modified albumin by AGEs may be more resistant to enzymatic degradation, which further increases the local accumulation of AGEs in tissues. The aim of the present study was to analyze in vitro glycation of serum albumin in the presence of piracetam (PIR) and the gliclazide (GLZ)-glycated albumin interaction. The analysis of PIR as an inhibitor and GLZ interaction with nonglycated human albumin (HSA) and glycated by fructose human albumin (gHSAFRC), in the absence and presence of piracetam (gHSAFRC-PIR), was performed by fluorescence quenching of macromolecules. On the basis of obtained data we concluded that under the influence of glycation, association constant ( K a ) of gliclazide to human serum albumin decreases and GLZ binds to HSA with less strength than under physiological conditions. PIR strongly inhibited the formation of AGEs in the system where the efficiency of HSA glycation was the largest. The analysis of piracetam influence on the GLZ-glycated albumin interaction has shown that piracetam increases the binding strength of GLZ to glycated albumin and weakens its therapeutic effect. Based on the obtained data we concluded that monitoring therapy and precautions are required in the treatment when the combinations of gliclazide and piracetam are used at the same time.

scholarly journals Differential Association of Plasmodium falciparum Na+/H+Exchanger Polymorphism and Quinine Responses in Field- and Culture-Adapted Isolates of Plasmodium falciparum

2011 ◽  
Vol 55 (12) ◽  
pp. 5834-5841 ◽  
Author(s):  
Stéphane Pelleau ◽  
Lionel Bertaux ◽  
Sébastien Briolant ◽  
Michael T. Ferdig ◽  
Véronique Sinou ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Drug Testing ◽  
Inhibitory Concentration ◽  
Cumulative Effect ◽  
Differential Association ◽  
Microsatellite Sequence ◽  
Content Type ◽  
Field Isolates ◽  
Selection Of

ABSTRACTPlasmodium falciparumisolates with decreased susceptibility to quinine are increasingly being found in malaria patients. Mechanisms involved in this resistance are not yet understood. Several studies claim that alongside mutations in the Pfcrtand Pfmdr1genes, the Pfnhe-1Na+/H+exchanger polymorphism plays a role in decreasing susceptibility. However, conflicting results on the link between the Pfnhe-1gene and quinine resistance arise from field- and culture-adapted isolates. We tested the association between Pfnhe-1, Pfcrt, and Pfmdr1polymorphisms in field- and culture-adapted isolates from various countries with theirin vitrosusceptibility to quinine. Field isolates presented a higher diversity of the Pfnhe-1microsatellite sequence than culture-adapted isolates. In culture-adapted isolates but not in field isolates, mutations in the Pfcrtand Pfmdr1genes, as well as a higher number of DNNND repeats in the Pfnhe-1gene, were associated with a higher 50% inhibitory concentration (IC50) of quinine. Furthermore, most of the culture-adapted isolates with more than one DNNND repeat in the Pfnhe-1gene also harbored mutated Pfcrtand Pfmdr1genes with an apparent cumulative effect on quinine susceptibility. This study supports the involvement of the Pfnhe-1gene in the modulation of thein vitroquinine response when associated with mutated Pfcrtand Pfmdr1genes. Culture adaptation could be responsible for selection of specific haplotypes of these three genes. Methods used for drug testing might thus influence the association between Pfnhe-1polymorphism and quinine susceptibility. However, we do not exclude the possibility that in particular settings, Pfnhe-1polymorphism can be used as a molecular marker for surveillance of quinine resistance.

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