n-Alkane and long-chain alcohol recovery in moose (Alces alces), a browsing herbivore

Habitat management for herbivores often depends on an understanding of the food habits of animals. Plant cuticular waxes containing nearly indigestible complex mixture of n-alkanes and long-chain alcohols (LCOHs) have recently shown promise for diet analyses, but the accuracy of the technique depends strongly on the efficiency of recovery of the markers in feces. Fecal recovery of n-alkanes and LCOHs from 10 browse stems or leaves and two ensiled grass hays fed to moose (Alces alces (Linnaeus, 1758)) during in vivo digestion trials was investigated. n-Alkanes and LCOHs were extracted using a single-step accelerated solvent extraction technique and the recovery of these cuticular components was calculated from the feces of the animals. n-Alkane recoveries from feces averaged 0.82, ranging from a low of 0.58 (haylage) to a high of 0.95 (browse stems). LCOH recoveries averaged 0.92 across all forages, ranging from 0.80 (haylage) to a high of 1.13 (browse stems). n-Alkane and LCOH fecal recovery increased with increasing chain length, similar to findings in other studies. Although fecal recovery of n-alkanes and LCOHs were variable, we conclude that they are inversely related to forage digestibility, are consistent within forage classes, and are therefore predictable markers for use in assessing herbivore diets.

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Quality-Related Properties of Equine Immunoglobulins Purified by Different Approaches

Toxins ◽

10.3390/toxins12120798 ◽

2020 ◽

Vol 12 (12) ◽

pp. 798

Author(s):

Sanja Mateljak Lukačević ◽

Tihana Kurtović ◽

Maja Lang Balija ◽

Marija Brgles ◽

Stephanie Steinberger ◽

...

Keyword(s):

Protective Efficacy ◽

Complex Mixture ◽

Downstream Processing ◽

Exchange Chromatography ◽

Single Step ◽

Relative Distribution ◽

Igg Subclass ◽

Processing Strategies ◽

Horse Plasma

Whole IgG antivenoms are prepared from hyperimmune animal plasma by various refinement strategies. The ones most commonly used at industrial scale are precipitation by sodium or ammonium sulphate (ASP), and caprylic acid precipitation (CAP) of non-immunoglobulin proteins. The additional procedures, which have so far been used for experimental purposes only, are anion-exchange (AEX) and cation-exchange chromatography (CEX), as well as affinity chromatography (AC) using IgG’s Fc-binding ligands. These protocols extract the whole IgG fraction from plasma, which contains both venom-specific and therapeutically irrelevant antibodies. Such preparations represent a complex mixture of various IgG subclasses whose functional and/or structural properties, as well as relative distribution, might be affected differently, depending on employed purification procedure. The aim of this work was to compare the influence of aforementioned refinement strategies on the IgG subclass distribution, venom-specific protective efficacy, thermal stability, aggregate formation and retained impurity profile of the final products. A unique sample of Vipera ammodytes ammodytes specific hyperimmune horse plasma was used as a starting material, enabling direct comparison of five purification approaches. The highest purity was achieved by CAP and AC (above 90% in a single step), while the lowest aggregate content was present in samples from AEX processing. Albumin was the main contaminant in IgG preparations obtained by ASP and CEX, while transferrin dominantly contaminated IgG sample from AEX processing. Alpha-1B-glycoprotein was present in CAP IgG fraction, as well as in those from ASP- and AEX-based procedures. AC approach induced the highest loss of IgG(T) subclass. CEX and AEX showed the same tendency, while CAP and ASP had almost no impact on subclass distribution. The shift in IgG subclass composition influenced the specific protective efficacy of the respective final preparation as measured in vivo. AC and CEX remarkably affected drug’s venom-neutralization activity, in contrary to the CAP procedure, that preserved protective efficacy of the IgG fraction. Presented data might improve the process of designing and establishing novel downstream processing strategies and give guidance for optimization of the current ones by providing information on potency-protecting and purity-increasing properties of each purification principle.

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Development of Single-step Protocols for the Separation of Naphtodianthrones from St. John�s Wort Extracts

Revista de Chimie ◽

10.37358/rc.18.8.6519 ◽

2018 ◽

Vol 69 (8) ◽

pp. 2295-2299

Author(s):

Elena Ionescu ◽

Tanta Verona Iordache ◽

Carmen Elena Tebrencu ◽

Ruxandra Mihaela Cretu ◽

Ana Mihaela Florea ◽

...

Keyword(s):

Hypericum Perforatum ◽

Single Step ◽

Anti Cancer ◽

Hypericum Perforatum L

St. John s Wort (SJW) or Hypericum perforatum L. is a therapeutic plant highly used in pharmacology. Recent in vivo anti-cancer action of naphtodianthrones (NTs) has extended the research related to enrichment methodologies of SJW phyto-extracts. Therefore, the presented study pursuits the optimization of single-step extraction methodologies to obtain NTs-rich extracts from SJW.

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Glucose Oligosaccharide and Long-Chain Glucomannan Feed Additives Induce Enhanced Activation of Intraepithelial NK Cells and Relative Abundance of Commensal Lactic Acid Bacteria in Broiler Chickens

Veterinary Sciences ◽

10.3390/vetsci8060110 ◽

2021 ◽

Vol 8 (6) ◽

pp. 110

Author(s):

Nathalie Meijerink ◽

Jean E. de Oliveira ◽

Daphne A. van Haarlem ◽

Guilherme Hosotani ◽

David M. Lamot ◽

...

Keyword(s):

Immune System ◽

Nk Cells ◽

Relative Abundance ◽

Nk Cell ◽

Feed Additives ◽

Microbiota Composition ◽

In Ovo ◽

Long Chain

Restrictions on the use of antibiotics in the poultry industry stimulate the development of alternative nutritional solutions to maintain or improve poultry health. This requires more insight in the modulatory effects of feed additives on the immune system and microbiota composition. Compounds known to influence the innate immune system and microbiota composition were selected and screened in vitro, in ovo, and in vivo. Among all compounds, 57 enhanced NK cell activation, 56 increased phagocytosis, and 22 increased NO production of the macrophage cell line HD11 in vitro. Based on these results, availability and regulatory status, six compounds were selected for further analysis. None of these compounds showed negative effects on growth, hatchability, and feed conversion in in ovo and in vivo studies. Based on the most interesting numerical results and highest future potential feasibility, two compounds were analyzed further. Administration of glucose oligosaccharide and long-chain glucomannan in vivo both enhanced activation of intraepithelial NK cells and led to increased relative abundance of lactic acid bacteria (LAB) amongst ileum and ceca microbiota after seven days of supplementation. Positive correlations between NK cell subsets and activation, and relative abundance of LAB suggest the involvement of microbiota in the modulation of the function of intraepithelial NK cells. This study identifies glucose oligosaccharide and long-chain glucomannan supplementation as effective nutritional strategies to modulate the intestinal microbiota composition and strengthen the intraepithelial innate immune system.

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A Novel Mouse Monoclonal Antibody C42 against C-Terminal Peptide of Alpha-1-Antitrypsin

International Journal of Molecular Sciences ◽

10.3390/ijms22042141 ◽

2021 ◽

Vol 22 (4) ◽

pp. 2141

Author(s):

Srinu Tumpara ◽

Elena Korenbaum ◽

Mark Kühnel ◽

Danny Jonigk ◽

Beata Olejnicka ◽

...

Keyword(s):

Monoclonal Antibody ◽

Flow Cytometry ◽

Western Blotting ◽

Complex Mixture ◽

Immunoglobulin M ◽

Confocal Laser ◽

Enzyme Linked Immunosorbent Assay ◽

Dot Blot

The C-terminal-fragments of alpha1-antitrypsin (AAT) have been identified and their diverse biological roles have been reported in vitro and in vivo. These findings prompted us to develop a monoclonal antibody that specifically recognizes C-36 peptide (corresponding to residues 359–394) resulting from the protease-associated cleavage of AAT. The C-36-targeting mouse monoclonal Immunoglobulin M (IgM) antibody (containing κ light chains, clone C42) was generated and enzyme-linked immunosorbent assay (ELISA)-tested by Davids Biotechnologie GmbH, Germany. Here, we addressed the effectiveness of the novel C42 antibody in different immunoassay formats, such as dot- and Western blotting, confocal laser microscopy, and flow cytometry. According to the dot-blot results, our novel C42 antibody detects the C-36 peptide at a range of 0.1–0.05 µg and shows no cross-reactivity with native, polymerized, or oxidized forms of full-length AAT, the AAT-elastase complex mixture, as well as with shorter C-terminal fragments of AAT. However, the C42 antibody does not detect denatured peptide in SDS-PAGE/Western blotting assays. On the other hand, our C42 antibody, unconjugated as well as conjugated to DyLight488 fluorophore, when applied for immunofluorescence microscopy and flow cytometry assays, specifically detected the C-36 peptide in human blood cells. Altogether, we demonstrate that our novel C42 antibody successfully recognizes the C-36 peptide of AAT in a number of immunoassays and has potential to become an important tool in AAT-related studies.

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Effect of long-chain fatty acyl-CoA on mitochondrial and cytosolic ATP/ADP ratios in the intact liver cell

Biochemical Journal ◽

10.1042/bj2200371 ◽

1984 ◽

Vol 220 (2) ◽

pp. 371-376 ◽

Cited By ~ 40

Author(s):

S Soboll ◽

H J Seitz ◽

H Sies ◽

B Ziegler ◽

R Scholz

Keyword(s):

Liver Cell ◽

Adenine Nucleotide ◽

Intact Cell ◽

Inhibitory Effect ◽

Fatty Acyl ◽

Fed State ◽

Physiological Relevance ◽

Chain Acyl ◽

Long Chain

The effect of long-chain acyl-CoA on subcellular adenine nucleotide systems was studied in the intact liver cell. Long-chain acyl-CoA content was varied by varying the nutritional state (fed and starved states) or by addition of oleate. Starvation led to an increase in the mitochondrial and a decrease in the cytosolic ATP/ADP ratio in liver both in vivo and in the isolated perfused organ as compared with the fed state. The changes were reversed on re-feeding glucose in liver in vivo or on infusion of substrates (glucose, glycerol) in the perfused liver, respectively. Similar changes in mitochondrial and cytosolic ATP/ADP ratios occurred on addition of oleate, but, importantly, not with a short-chain fatty acid such as octanoate. It is concluded that long-chain acyl-CoA exerts an inhibitory effect on mitochondrial adenine nucleotide translocation in the intact cell, as was previously postulated in the literature from data obtained with isolated mitochondria. The physiological relevance with respect to pyruvate metabolism, i.e. regulation of pyruvate carboxylase and pyruvate dehydrogenase by the mitochondrial ATP/ADP ratio, is discussed.

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Formycin B-resistant mutants of Chinese hamster ovary cells: novel genetic and biochemical phenotype affecting adenosine kinase

Molecular and Cellular Biology ◽

10.1128/mcb.3.8.1468-1477.1983 ◽

1983 ◽

Vol 3 (8) ◽

pp. 1468-1477

Author(s):

K D Mehta ◽

R S Gupta

Keyword(s):

Chinese Hamster Ovary ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Complex Form ◽

Single Step ◽

Adenosine Kinase ◽

Cross Resistance ◽

Ovary Cells ◽

Cell Extracts

Stable mutants which are approximately three- and eightfold resistant to the pyrazolopyrimidine nucleosides formycin A and formycin B (FomR) have been selected in a single step from mutagenized Chinese hamster ovary cells. In cell extracts, the two FomR mutants which were examined were both found to contain no measurable activity of the enzyme adenosine kinase (AK). However, cross-resistance studies with other adenosine analogs such as toyocamycin and tubercidin show that these mutants are distinct from toyocamycin or tubercidin resistant (Toyr) mutants which also contain no measurable AK activity in cell extracts. Studies on the uptake and incorporation of [3H]adenosine and [3H]tubercidin by various mutants and parental cell lines show that unlike the Toyr mutants, which are severely deficient in the phosphorylation of these compounds, the FomR mutants possess nearly normal capacity to phosphorylate these compounds and incorporate them into cellular macromolecules. These results suggest that the FomR mutants contain normal levels of AK activity in vivo. In cell hybrids formed between FomR X FomS cells and FomR X Toyr cells, the formycin-resistant phenotype of both of the FomR mutants behaved codominantly. However, the extracts from these hybrid cells contained either congruent to 50% (FomR X FomS) or no measurable (FomR X Toyr) AK activity, indicating that the lesion in these mutants neither suppresses the wild-type AK activity nor complements the AK deficiency of the Toyr mutants. The presence of AK activity in the FomR mutants in vivo, but not in their cell extracts, along with the codominant behavior of the mutants in hybrids, indicates that the lesions in the FomR mutant are of a novel nature. It is suggested that the genetic lesion in these mutants affects AK activity indirectly and that it may involve an essential cellular function which exists in a complex form with AK. Some implications of these results regarding the mechanism of action of formycin B are discussed.

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Long-Chain Fatty Acid Transport at the Blood-Brain Barrier and Incorporation into Brain Phospholipids: A New In Vivo Method for Examining Neuroplasticity, and Brain Second Messenger Systems Involving Phospholipase A2 Activation

New Concepts of a Blood—Brain Barrier ◽

10.1007/978-1-4899-1054-7_13 ◽

1995 ◽

pp. 119-140 ◽

Cited By ~ 4

Author(s):

S. I. Rapoport ◽

P. J. Robinson

Keyword(s):

Fatty Acid ◽

Phospholipase A2 ◽

Blood Brain Barrier ◽

Second Messenger ◽

Chain Fatty Acid ◽

Fatty Acid Transport ◽

Acid Transport ◽

Long Chain Fatty Acid ◽

Long Chain

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Factor XII Activation Promotes Platelet Consumption in the Presence of Bacterial-Type Long-Chain Polyphosphate In Vitro and In Vivo

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvbaha.118.311193 ◽

2018 ◽

Vol 38 (8) ◽

pp. 1748-1760 ◽

Cited By ~ 15

Author(s):

Jevgenia Zilberman-Rudenko ◽

Stéphanie E. Reitsma ◽

Cristina Puy ◽

Rachel A. Rigg ◽

Stephanie A. Smith ◽

...

Keyword(s):

Factor Xii ◽

Bacterial Type ◽

Long Chain

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Ex VivoMaturation Assay for Testing Antimalarial Sensitivity of Rodent Malaria Parasites

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01292-16 ◽

2016 ◽

Vol 60 (11) ◽

pp. 6859-6866 ◽

Cited By ~ 4

Author(s):

Zi Wei Chang ◽

Benoit Malleret ◽

Bruce Russell ◽

Laurent Rénia ◽

Carla Claser

Keyword(s):

Ex Vivo ◽

Single Step ◽

Parasite Development ◽

Ring Stage ◽

Malaria Parasites ◽

Rodent Malaria ◽

Content Type ◽

Parasite Biology

ABSTRACTEx vivoassay systems provide a powerful approach to studying human malaria parasite biology and to testing antimalarials. For rodent malaria parasites, short-termin vitroculture andex vivoantimalarial susceptibility assays are relatively cumbersome, relying onin vivopassage for synchronization, since ring-stage parasites are an essential starting material. Here, we describe a new approach based on the enrichment of ring-stagePlasmodium berghei,P. yoelii, andP. vinckei vinckeiusing a single-step Percoll gradient. Importantly, we demonstrate that the enriched ring-stage parasites develop synchronously regardless of the parasite strain or species used. Using a flow cytometry assay with Hoechst and ethidium or MitoTracker dye, we show that parasite development is easily and rapidly monitored. Finally, we demonstrate that this approach can be used to screen antimalarial drugs.

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