Determination of Angiotensin-(1–7) with HPLC/Fluorescence-Detection

Abstract

Angiotensin-(1-7) is an important active component in the renin-angiotensin-system. Due to its cardio protective effects it is now under investigation in combination with antioxidants as a reperfusion solution. The combination showed impressive effects on isolated hearts of male Wistar rats after induced ischemia. In this work a high performance liquid chromatography method with fluorescence detection was developed for the first time for in-process measurements as well as for stability tests of the peptide in the novel antioxidant-containing Karal® solution. For fluorescence detection of angiotensin-(1-7) fluorescamine as derivatization dye was applied. Under optimized conditions the method showed linearity over the range of 50 to 5000 ng/mL with R(2) of 0.9988 and an overall precision better than 5.0 %. LOD and LOQ were determined to be in the femtomol range on column. It was found that stability of angiotensin-(1-7) could be significantly improved in the antioxidant containing preparation compared to aqueous solutions.

Stabilization of Angiotensin-(1-7) in Cardioprotective Solutions

“…HPLC analysis was carried out with an Agilent Technologies 1260 Infinity Quat Pump VL and a RF-10AXL Shimadzu Fluorescence Detector. Quantification was performed as described elsewhere (Russ et al 2016). Under these conditions the retention time of Ang 1-7 was about 4.7 min.…”

“…LOD was determined to be 15 ng/mL. The peptide was derivatized by adding an appropriate amount of fluorescamine at room temperature, likewise according to Russ et al (2016). The reaction mixture was then injected into the HPLC system via a 20 µL loop and detected at 390 nm excitation and 470 nm emission wavelength.…”

“…An HPLC method with fluorescence detection was used to study the stability of Ang 1-7 in aqueous solutions (Russ et al 2016), while offline nano-ESI MS was also applied to detect possible degradation products with high sensitivity and specificity.…”

“…Both used a water/acetonitrile mixture with formic acid as solvent for the Ang 1-7 solution, but without information on storage conditions, pH value and additives (Cui et al 2007;Ali et al 2014). During the development of our HPLC method for determining Ang 1-7 (Russ et al 2016), a stability enhancement after addition of an antioxidative solution was noticed. However, up to now no information is given on causes of such an enhancement and possibilities for further improvement.…”

Using non-empirically tuned range-separated functionals with simulated emission bands to model fluorescence lifetimes

“…Other applications of fluorescence spectroscopy and microscopy can be found in molecular sensing, [6][7][8] biomedical imaging, 9,10 and chromatographic detectors. [11][12][13] Properties that are desirable for a fluorophore include a high molar absorptivity and fluorescence quantum yield, resistance towards photobleaching, good solubility in various solvents, and absorption and emission wavelengths that are suited towards the application of interest. However, fluorophores with an optimal performance in all aspects for a particular application are rarely found.…”