Cation exchange HPLC analysis of desmosines in elastin hydrolysates

Abstract

Desmosine crosslinks are responsible for the elastic properties of connective tissues in lungs and cardiovascular system and are often compromised in disease states. We developed a new, fast, and simple cation exchange HPLC assay for the analysis of desmosine and isodesmosine in animal elastin. The method was validated by determining linearity, accuracy, precision, and desmosines stability and was applied to measure levels of desmosines in porcine and murine organs. The detection and quantification limits were 2 and 4 pmol, respectively. The run-time was 8 min. Our cation exchange column does not separate desmosine and isodesmosine, but their level can be quantified from absorbance at different wavelengths. Using this assay, we found that desmosines levels were significantly lower in elastin isolated from various organs of immunodeficient severe combined immunodeficiency mice compared with wild-type animals. We also found that desmosines levels were lower in lung elastin isolated from hyperhomocysteinemic Pcft −/− mice deficient in intestinal folate transport compared with wild-type Pcft +/+ animals.Electronic Supplementary MaterialSupplementary material is available for this article at 10.1007/s00216-011-5346-z and is accessible for authorized users.

Towards rebuilding vaginal support utilizing an extracellular matrix bioscaffold

“…Mature elastin content was measured by ultrahigh performance liquid chromography (UPLC) as previously described [39, 40]. Briefly, hydrolyzed tissue was applied to Oasis MCX cation exchange SPE columns (Waters, Milford Massachusetts), and eluted with ammonium hydroxide/water/methanol (10/40/50).…”

N‐Homocysteinylation impairs collagen cross‐linking in cystathionine β‐synthase‐deficient mice: a novel mechanism of connective tissue abnormalities

“…Collagen Pyd cross‐links were quantified by HPLC (24). Collagen was acid hydrolyzed (6 N HCl; 110°C; 24 h), and Pyds were separated on a cation exchange polysulfoethyl aspartamide column, 100 × 2.1 mm, 3 μM, 300 A (Poly LC), with a salt gradient at flow rate of 0.36 ml/min at 24°C.…”

Micellar Electrokinetic Chromatography with Laser Induced Detection and liquid chromatography tandem mass-spectrometry-based desmosine assays in urine of patients with Chronic Obstructive Pulmonary Disease: A comparative analysis

“…On the other hand, the high number of different techniques developed by several laboratories around the world for their screening and quantification in a variety of fluids, reflects a growing interest in Des as an end-point in clinical trials. These techniques span from Radio Immuno Assay (RIA) [8][9][10][11] to High Performance Liquid Chromatography (HPLC) [12][13][14][15][16] and capillary electrophoresis (CE) [17]. In particular, the high sensitivity of Micellar Electrokinetic Chromatography with Laser Induced Fluorescence detection (MEKC-LIF), has proved successful in detecting these cross-links for the first time in serum and sputum as well [18].…”