Measuring urinary cortisol and testosterone levels in male Barbary macaques: A comparison of EIA and LC-MS

The development of methods to quantify hormones from non-invasively collected samples such as urine or feces has facilitated endocrinology research on wild-living animals. To ensure that hormone measurements are biologically meaningful, method validations are highly recommended for each new species or sample matrix. Our aim was to validate three commonly used enzyme immunoassays (EIA), one for analysis of cortisol and two for analysis of testosterone, to assess adrenocortical and gonadal activity, respectively, from the urine of male Barbary macaques. We compared EIA and liquid chromatography--mass spectrometry (LC-MS) results to determine if the EIA measurements truly reflect levels of the target hormone and to determine if cross-reactivities with other steroids were potentially confounding results. Furthermore, we conducted a biological validation of testosterone to ensure that both EIA and LC-MS were able to capture physiologically meaningful differences in hormone levels. We found that cortisol measured by EIA correlated strongly with cortisol measured by LC-MS in both adult and immature males, without the need for deconjugation of steroids in the urine. Both testosterone EIAs correlated strongly with LC-MS in adult males, but only if steroids in the urine were deconjugated by enzymatic hydrolysis prior to analysis. However, in immature males, EIA and LC-MS results did not correlate significantly. Further correlation analyses suggest this is likely due to cross-reactivity of the testosterone antibody with other adrenal steroids such as cortisol, DHEA, and likely others, which are present at much higher concentrations relative to testosterone in immature males. Testosterone levels were significantly higher in adult compared to immature males as measured by LC-MS but not as measured by EIA. Taken together, our results suggest that the testosterone EIAs are suitable to assess gonadal activity in adult but not immature males, and only if a hydrolysis of the urine is conducted prior to analysis.

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Measuring urinary cortisol and testosterone levels in male Barbary macaques: A comparison of EIA and LC–MS

General and Comparative Endocrinology ◽

10.1016/j.ygcen.2019.05.017 ◽

2019 ◽

Vol 281 ◽

pp. 117-125 ◽

Cited By ~ 2

Author(s):

Alan V. Rincon ◽

Julia Ostner ◽

Michael Heistermann ◽

Tobias Deschner

Keyword(s):

Urinary Cortisol ◽

Barbary Macaques ◽

Testosterone Levels

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HIV-Infected Patients: Cross Site-Specific Hydrolysis of H3 and H4 Histones and Myelin Basic Protein with Antibodies against These Three Proteins

Molecules ◽

10.3390/molecules26020316 ◽

2021 ◽

Vol 26 (2) ◽

pp. 316

Author(s):

Svetlana V. Baranova ◽

Pavel S. Dmitrenok ◽

Valentina N. Buneva ◽

Sergey E. Sedykh ◽

Georgy A. Nevinsky

Keyword(s):

Myelin Basic Protein ◽

Intercellular Space ◽

Basic Protein ◽

Cross Reactivity ◽

Cleavage Sites ◽

Site Specific ◽

Protein Clusters ◽

Negative Role ◽

Toxic Responses ◽

Hydrolysis Of

Histones play important roles in chromatin functioning and gene transcription, but in the intercellular space, they are harmful since they stimulate systemic inflammatory and toxic responses. Electrophoretically homogeneous IgGs against myelin basic protein (MBP), as well as H3 and H4 histones, were isolated from sera of HIV-infected patients. In contrast to known classical proteases, these IgGs split exclusively only histones and MBP but no other control proteins. Among 13 sites of hydrolysis of H3 by IgGs against H3 and 14 sites for anti-MBP IgGs, only two sites of the hydrolysis were the same. Between seven cleavage sites of H4 with IgGs against H4 and 9 sites of this histone hydrolysis by antibodies against MBP, only three sites were the same. The sites of hydrolysis of H3 (and H4) with abzymes against these histones and against MBP were different, but several expended protein clusters containing hydrolysis sites are partially overlapped. The existence of enzymatic cross-reactivity of abzymes against H3 and H4 and MBP represents a great menace to humans since due to cell apoptosis, histones constantly occur in human blood. They can hydrolyze MBP of the myelin sheath of axons and play a negative role in the pathogenesis of HIV-infected patients.

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Targeted analysis of phenolic compounds by LC-MS v1

10.17504/protocols.io.byhcpt2w ◽

2021 ◽

Author(s):

Bashar Amer ◽

Ramu Kakumanu ◽

yangtian not provided ◽

Aymerick Eudes ◽

Edward EK Baidoo

Keyword(s):

Phenolic Compounds ◽

Plant Biomass ◽

Reversed Phase ◽

Industrial Applications ◽

Liquid Chromatography Mass Spectrometry ◽

Reversed Phase Liquid Chromatography ◽

Sample Matrix ◽

Targeted Analysis ◽

Challenges And Opportunities ◽

Phase Liquid

Cell-wall-bound (CWB) aromatics such as ferulate and p-coumarate play important physiological roles in plant development and response to stresses. Their presence also poses some challenges and opportunities during processing of plant biomass in various agro-industrial applications. To this end, we have developed a robust high-throughput reversed-phase liquid chromatography mass spectrometry method for quantifying CWB phenolic compounds. The method showed excellent linearity (R2 = ≥0.999) and intraday retention time repeatability (≤ 0.31 %RSD) for ferulate and p-coumarate. The limits of detection and quantitation for these analytes were ≤ 39 nM and 130 nM, respectively. Furthermore, there was very little effect of the CWB sample matrix on the retention times of the analytes and analyte percent recoveries from the CWB sample matrix was ≥83.91%.

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Not only big bulls. Correlation between morphometry, reproductive success, and testosterone level in a flooded savannah population of the spectacled caiman (Caiman crocodilus)

Canadian Journal of Zoology ◽

10.1139/cjz-2020-0099 ◽

2021 ◽

Author(s):

Leidy Alejandra Barragan Contreras ◽

Rafael Antelo ◽

Adolfo Amezquita

Keyword(s):

Reproductive Success ◽

Breeding Season ◽

Steroid Hormone ◽

Multiple Paternity ◽

Adult Males ◽

Behavioral Traits ◽

Testosterone Levels ◽

Caiman Crocodilus ◽

Paternity Tests ◽

Hourly Basis

Testosterone is a steroid hormone involved in the expression of many morphological, physiological, and behavioral traits that arguably affect reproductive success. The evidence for that link is, however, incomplete or absent in the research on crocodylian species. Testosterone levels are also known to change throughout the breeding season, often on an hourly basis, which may further complicate studying their relationship with breeding success. We tested here whether baseline testosterone levels, measured out of the breeding season, are correlated with morphometry and reproductive success in Caiman crocodilus (LINNAEUS, 1758). Paternity tests, based on the amplification and genotyping of eight fluorochrome labeled microsatellites, failed to support a continuous relationship between these variables. Although adult males of all sizes contribute to reproduction, paternity was overrepresented in a few males with high values of maleness index (bigger males), supporting a despotic or pyramidal hierarchy among males. Maternity assignments supported the existence of multiple paternity, a phenomenon previously attributed in this species to the lack of large males caused by human hunting. The idea of larger males having more offspring is widespread in crocodylians, but to our knowledge, this is the first investigation that prove this dogma.

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New insights into lactase and glycosylceramidase activities of rat lactase-phlorizin hydrolase

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1989.257.4.g616 ◽

1989 ◽

Vol 257 (4) ◽

pp. G616-G623 ◽

Cited By ~ 2

Author(s):

H. A. Buller ◽

A. G. Van Wassenaer ◽

S. Raghavan ◽

R. K. Montgomery ◽

M. A. Sybicki ◽

...

Keyword(s):

Active Sites ◽

Specific Activity ◽

Fold Increase ◽

Developmental Pattern ◽

Lactose Hydrolysis ◽

Adult Males ◽

Lactase Activity ◽

Developmental Patterns ◽

Catalytic Sites ◽

Hydrolysis Of

Lactase-phlorizin hydrolase, a small intestinal disaccharidase, has been considered mainly an enzyme important only for the hydrolysis of lactose. After weaning in most mammals lactase-specific activity falls markedly, and, functionally, adult mammals are considered to be lactase deficient. However, the persistence of low levels of lactase activity in adulthood has never been explained. In addition, it has been suggested that lactase-phlorizin hydrolase is associated with glycosylceramidase activity when the enzyme is prepared by column chromatography, but it is unclear whether this represents copurified activities or two catalytic sites on one peptide. The developmental patterns of lactase-phlorizin hydrolase and other disaccharidases were investigated in homogenates of total rat small intestine; lactase and several glycosylceramidases were measured in immunoprecipitates from these homogenates using a monoclonal antibody. The developmental pattern of total lactase activity showed a steady 2.3-fold increase to adult levels (specific activity decreased eightfold), whereas total phlorizin-hydrolase activity increased 10.7-fold (specific activity decreased threefold). As expected, levels of both total and specific sucrase and maltase activities increased during development. In lactating rats total lactase activity showed a significant increase compared with adult males. The developmental pattern of the enzyme activities for the glycolipid substrates was similar to that found for lactase, and the immunoprecipitated enzyme showed a 40- to 55-fold higher affinity for the glycolipids than for lactose. Galactosyl- and lactosylceramide inhibited lactose hydrolysis by 38%, without a competitive pattern, suggesting two different active sites for lactose and glycolipid hydrolysis, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

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Flow Chemistry System for Carbohydrate Analysis by Rapid Labeling of Saccharides after Glycan Hydrolysis

SLAS TECHNOLOGY Translating Life Sciences Innovation ◽

10.1177/2472630320924620 ◽

2020 ◽

Vol 25 (4) ◽

pp. 356-366

Author(s):

Wei-Ting Hung ◽

Yi-Ting Chen ◽

Chung-Hsuan Chen ◽

Yuan Chuan Lee ◽

Jim-Min Fang ◽

...

Keyword(s):

Mass Spectrometry ◽

Flow System ◽

Flow Chemistry ◽

Composition Analysis ◽

Laser Desorption Ionization ◽

Liquid Chromatography Mass Spectrometry ◽

Ionization Time ◽

Flight Mass Spectrometry ◽

Rapid Labeling ◽

Hydrolysis Of

This study demonstrates the utilization of a flow chemistry system for continuous glycan hydrolysis and saccharide labeling to assist with the existing methods in glycan structural analysis. Acidic hydrolysis of glycans could be accelerated in a flow system. Aldoses and α-ketoacid-type saccharides were effectively labeled with naphthalene-2,3-diamine (NADA) at 60 °C for 10 min to form the fluorescent naphthimidazole (NAIM) and quinoxalinone (QXO) derivatives, respectively. The NADA-labeled derivatives improved the structural determination and composition analysis for their parent saccharides by using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), liquid chromatography mass spectrometry (LC-MS), and nuclear magnetic resonance (NMR). Furthermore, this protocol was applied to determine the SA–Gal–Glc sequence of GM3-sugar out of six possible permutations.

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Association between Urinary Triclosan and Serum Testosterone Levels in U.S. Adult Males from NHANES, 2011–2012

International Journal of Environmental Research and Public Health ◽

10.3390/ijerph17207412 ◽

2020 ◽

Vol 17 (20) ◽

pp. 7412 ◽

Cited By ~ 1

Author(s):

Judy Yan ◽

Michael A. Joseph ◽

Simone A. Reynolds ◽

Laura A. Geer

Keyword(s):

Endocrine Disruptor ◽

Linear Regression Analysis ◽

Serum Testosterone ◽

Adult Males ◽

Comprehensive Understanding ◽

Testosterone Levels ◽

Adult Men ◽

Health And Nutrition ◽

Multivariable Linear Regression Analysis ◽

Over Time

Triclosan was introduced into the market in the 1970s and has since been used as an antimicrobial agent in a diverse array of consumer and personal care products. Although it has been widely used over a number of years, there is growing concern and debate over its safety and efficacy and its potential as an endocrine disruptor. Although prior animal toxicology studies have shown an association between triclosan and decreased testosterone levels, human studies have been limited, particularly for adult men. Using the National Health and Nutrition Examination Survey data (NHANES, 2011–2012), we examined the association of urinary triclosan on testosterone levels in adult men 18–65 years of age. Multivariable linear regression analysis failed to show an association between triclosan and serum testosterone (β = 0.0003, p = 0.98, 95% CI = −0.024, 0.025). The results suggest there is no association or that triclosan concentrations are too low to cause a significant impact on testosterone levels. Additionally, longitudinal studies would provide a more comprehensive understanding of the direction of change and magnitude of causal relationships over time.

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Poor specificity and recovery of urinary free cortisol as determined by the Bayer ADVIA Centaur extraction method

Annals of Clinical Biochemistry International Journal of Laboratory Medicine ◽

10.1258/000456303322326506 ◽

2003 ◽

Vol 40 (5) ◽

pp. 563-565 ◽

Cited By ~ 9

Author(s):

G Gray ◽

L Shakerdi ◽

AM Wallace

Keyword(s):

Mass Spectrometry ◽

Gas Chromatography ◽

Solvent Extraction ◽

Extraction Method ◽

Urinary Free Cortisol ◽

Cross Reactivity ◽

Gas Chromatography Mass Spectrometry ◽

Sample Matrix ◽

Free Cortisol ◽

Free Serum

Background: Immunoassay methods for urinary free cortisol (UFC) lack specificity, and many procedures have not been fully evaluated for routine use. In the current study we evaluated the Bayer ADVIA Centaur extraction UFC immunoassay and compared results to those obtained by a specific gas chromatography-mass spectrometry (GC-MS) method. Results: The Bayer ADVIA Centaur cortisol assay lacked specificity. Cortisone, a steroid present in urine at concentrations similar to those of cortisol, demonstrated a cross-reactivity of 44%. In addition, the choice of matrix used to resuspend steroids after solvent extraction from urine affected recovery. Recovery was improved if the recommended urine reconstruction buffer was replaced by steroid-free serum. Conclusion: Irrespective of the sample matrix used, the Centaur method overestimates UFC, giving results up to twice those obtained by a specific GC-MS method.

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