{"id":43346,"date":"2020-10-20T14:30:00","date_gmt":"2020-10-20T14:30:00","guid":{"rendered":"https:\/\/discover.restek.com\/uncategorized\/simultaneous-analysis-of-catecholamines-and-metanephrines-in-urine-by-lc-msms\/"},"modified":"2025-12-15T13:32:23","modified_gmt":"2025-12-15T13:32:23","slug":"simultaneous-analysis-of-catecholamines-and-metanephrines-in-urine-by-lc-msms","status":"publish","type":"post","link":"https:\/\/discover.restek.com\/de\/application-notes\/cfan2800\/simultaneous-analysis-of-catecholamines-and-metanephrines-in-urine-by-lc-msms","title":{"rendered":"Simultaneous Analysis of Catecholamines and Metanephrines in Urine by LC-MS\/MS"},"content":{"rendered":"\n<h2 class=\"wp-block-heading\">Abstract<\/h2>\n\n\n\n<p>Clinical diagnosis of pheochromocytoma and paraganglioma is often based on the analysis of catecholamines (epinephrine, norepinephrine, dopamine) and metanephrines (metanephrine, normetanephrine, 3-methoxytyramine) in urine. Analysis of these polar compounds using reversed-phase LC can be difficult due to limited chromatographic retention, which results in poor separation of the analytes from closely eluting matrix interferences. This method overcomes these problems by combining a simple solid phase extraction procedure with the consistent and accurate chromatographic performance of a Raptor Biphenyl column.<\/p>\n\n\n\n<h2 class=\"wp-block-heading\">Introduction<\/h2>\n\n\n\n<p>Pheochromocytomas and paragangliomas are neuroendocrine tumors that are characterized by the release of elevated levels of catecholamines. The Endocrine Society, the American Association for Clinical Chemistry, and the European Society for Endocrinology have all released clinical practice guidelines for the diagnosis and management of these diseases and recommend two major diagnostic tests: (1) plasma free metanephrines, which is highly sensitive, and (2) 24-hour urinary collection of catecholamines and metanephrines, which is highly specific.<\/p>\n\n\n\n<p>The target analytes for urinary analysis are epinephrine (EPI), norepinephrine (NE), and dopamine (DA), as well as their respective methylated metabolites, metanephrine (MN), normetanephrine (NMN), and 3-methoxytyramine (3-MT) (Figure 1). Although reversed-phase LC-MS\/MS has been the method of choice for this analysis, challenges still remain because these polar analytes are difficult to retain and this, in combination with the presence of matrix interferences, can result in inconsistent chromatographic performance. To solve these problems, a simple and fast solid phase extraction (SPE) procedure was developed, followed by LC-MS\/MS analysis using a Raptor Biphenyl column. The method is accurate and robust for the simultaneous analysis of catecholamines and metanephrines in urine and, as such, is suitable for high-throughput clinical diagnostics.<\/p>\n\n\n<div class=\"wp-block-custom-chromatogram-article-top\"><div class=\"chromatogram-article-placeholder\"><div class=\"figure-heading\"><strong>Figure 1:<\/strong>\u00a0Structures of Catecholamines and Methylated Metabolites.<\/div><div class='chromatogram-article-inner-full'><div class=\"chromatogram-article-inner\">\n<style>.kb-image43346_6f8946-76 .kb-image-has-overlay:after{opacity:0.3;}<\/style>\n<div class=\"wp-block-kadence-image kb-image43346_6f8946-76\"><figure class=\"aligncenter size-large\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"548\" src=\"https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01-1024x548.jpg\" alt=\"\" class=\"kb-img wp-image-18904\" title=\"-\" srcset=\"https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01-1024x548.jpg 1024w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01-300x161.jpg 300w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01-768x411.jpg 768w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01-1536x823.jpg 1536w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-01.jpg 1800w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure><\/div>\n\n<\/div><\/div><\/div>\n\n\n<h2 class=\"wp-block-heading\">Experimental<\/h2>\n\n\n\n<h3 class=\"wp-block-heading\">Solid Phase Extraction<\/h3>\n\n\n\n<p>A 200 \u00b5L aliquot of urine sample was mixed with 10 \u03bcL of internal standard solution (1 \u00b5g\/mL in methanol) and 600 \u03bcL of 250 mM ammonium acetate solution. The mixture was loaded onto the EVOLUTE EXPRESS WCX 96-well plate (30 mg) and washed with 1 mL water and 1 mL methanol:acetonitrile (60:40). The elution was performed with 200 \u00b5L of water:methanol (95:5) solution containing 5% formic acid and then 2 \u03bcL was injected for analysis.<\/p>\n\n\n\n<h3 class=\"wp-block-heading\">Calibration Standards and Quality Control Samples<\/h3>\n\n\n\n<p>DC Mass Spect Gold Urine (Golden West Biologicals) was fortified with six analytes to prepare calibrated standards and QC samples. The linearity ranges were from 0.5-250 ng\/mL for epinephrine; 1-1000 ng\/mL for norepinephrine; 5-1500 ng\/mL for metanephrine and 3-methoxytyramine; and 10-2000 ng\/mL for dopamine and normetanephrine. Three QC levels were prepared at 2.5, 25, and 75 ng\/mL for epinephrine and norepinephrine; 25, 75, and 750 ng\/mL for metanephrine and 3-methoxytyramine; and 75, 750, and 1500 ng\/mL for dopamine and normetanephrine. The fortified standard and QC samples were subjected to the SPE procedure described above.<\/p>\n\n\n\n<h3 class=\"wp-block-heading\">Urine Analysis<\/h3>\n\n\n\n<p>To confirm that the method could accurately measure the urinary catecholamines and metabolites, two levels of urine samples (Bio-Rad Lyphochek quantitative urine controls) were analyzed with the established SPE and chromatographic methods. The normal urine was fortified with 10 ng\/mL of epinephrine, norepinephrine, 3-methoxytyramine, and 100 ng\/mL of dopamine, metanephrine, and normetanephrine. The abnormal urine was fortified with 50 ng\/mL of epinephrine and norepinephrine and 200 ng\/mL of dopamine, metanephrine, normetanephrine, 3-methoxytyramine. Accuracy was determined using the measured concentration difference between the blank and fortified urine.<\/p>\n\n\n\n<p>LC-MS\/MS analysis of catecholamines and metanephrines in urine was performed on an ACQUITY UPLC instrument coupled with a Waters Xevo TQ-S mass spectrometer. Instrument conditions were as follows and analyte transitions are provided in Table I.<\/p>\n\n\n<figure class=\"wp-block-table\">\n<table class=\"has-fixed-layout\" style=\"border-style: none;\">\n<tbody>\n<tr>\n<td>Analytical column:<\/td>\n<td colspan=\"2\">Raptor Biphenyl (2.7 \u00b5m, 150 mm x 2.1 mm; cat.#\u00a09309A62)<\/td>\n<\/tr>\n<tr>\n<td>Mobile phase A:<\/td>\n<td colspan=\"2\">0.2% Formic acid in water<\/td>\n<\/tr>\n<tr>\n<td>Mobile phase B:<\/td>\n<td colspan=\"2\">0.2% Formic acid in methanol<\/td>\n<\/tr>\n<tr>\n<td>Gradient:<\/td>\n<td>Time (min)<\/td>\n<td>%B<\/td>\n<\/tr>\n<tr>\n<td>\u00a0<\/td>\n<td>0.00<\/td>\n<td>5<\/td>\n<\/tr>\n<tr>\n<td>\u00a0<\/td>\n<td>2.50<\/td>\n<td>25<\/td>\n<\/tr>\n<tr>\n<td>\u00a0<\/td>\n<td>3.00<\/td>\n<td>95<\/td>\n<\/tr>\n<tr>\n<td>\u00a0<\/td>\n<td>3.01<\/td>\n<td>5<\/td>\n<\/tr>\n<tr>\n<td>\u00a0<\/td>\n<td>5.00<\/td>\n<td>5<\/td>\n<\/tr>\n<tr>\n<td>Flow rate:<\/td>\n<td colspan=\"2\">0.3 mL\/min<\/td>\n<\/tr>\n<tr>\n<td>Injection volume:<\/td>\n<td colspan=\"2\">2 \u00b5L<\/td>\n<\/tr>\n<tr>\n<td>Column temp.:<\/td>\n<td colspan=\"2\">30 \u00b0C<\/td>\n<\/tr>\n<tr>\n<td>Ion mode:<\/td>\n<td colspan=\"2\">Positive ESI<\/td>\n<\/tr>\n<\/tbody>\n<\/table>\n<\/figure>\n\n\n<p><strong>Table I:<\/strong>&nbsp;Analyte Transitions<\/p>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><tbody><tr><td><strong>Analyte<\/strong><\/td><td><strong>Precursor Ion<\/strong><\/td><td><strong>Product Ion<br>Quantifier<\/strong><\/td><td><strong>Product Ion<br>Qualifier<\/strong><\/td><\/tr><tr><td>Norepinephrine-d6<\/td><td>158.16<\/td><td>111.26<\/td><td>\u2014<\/td><\/tr><tr><td>Norepinephrine<\/td><td>152.07<\/td><td>106.91<\/td><td>134.97<\/td><\/tr><tr><td>Epinephrine-d6<\/td><td>172.16<\/td><td>111.86<\/td><td>\u2014<\/td><\/tr><tr><td>Epinephrine<\/td><td>166.07<\/td><td>135.00<\/td><td>107.02<\/td><\/tr><tr><td>Normetanephrine-d3<\/td><td>169.00<\/td><td>136.96<\/td><td>\u2014<\/td><\/tr><tr><td>Normetanephrine<\/td><td>166.00<\/td><td>148.99<\/td><td>134.02<\/td><\/tr><tr><td>Dopamine-d4<\/td><td>141.16<\/td><td>94.69<\/td><td>\u2014<\/td><\/tr><tr><td>Dopamine<\/td><td>136.97<\/td><td>64.66<\/td><td>91.09<\/td><\/tr><tr><td>Metanephrine-d3<\/td><td>183.00<\/td><td>151.15<\/td><td>\u2014<\/td><\/tr><tr><td>Metanephrine<\/td><td>179.94<\/td><td>165.01<\/td><td>148.22<\/td><\/tr><tr><td>3-Methoxytyramine-d4<\/td><td>155.07<\/td><td>122.93<\/td><td>\u2014<\/td><\/tr><tr><td>3-Methoxytyramine<\/td><td>151.00<\/td><td>91.02<\/td><td>65.05<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<h2 class=\"wp-block-heading\">Results and Discussion<\/h2>\n\n\n\n<h3 class=\"wp-block-heading\">Chromatographic Performance<\/h3>\n\n\n\n<p>The analysis of normal human urine (Bio-Rad Lyphochek quantitative urine control, level 1) demonstrates that a fast 5-minute chromatographic analysis is achieved with direct injection of the elution solution that was obtained from the simple SPE procedure (Figure 2). The Raptor Biphenyl column provided adequate retention such that all target analytes could be quantified with no observed influence from matrix interferences.<\/p>\n\n\n<div class=\"wp-block-custom-chromatogram-article-top\"><div class=\"chromatogram-article-placeholder\"><div class=\"figure-heading\"><strong>Figure 2:<\/strong>\u00a0Analysis of Catecholamines and Metanephrines in Urine.<\/div><div class='chromatogram-article-inner-full'><div class=\"chromatogram-article-inner\">\n<div class=\"wp-block-custom-chromatogram-article\"><div class=\"wp-block-custom-chromatogram-article\"><div class=\"chromatogram-image regular-image\"><img decoding=\"async\" src=\"https:\/\/ez.restek.com\/images\/cgram\/lc_cf0703.png\" alt=\"Catecholamines and Metabolites in Human Urine on Raptor Biphenyl by LC-MS\/MS\" title=\"-\"><\/div><p class=\"article-id\" style=\"text-align:center\"> LC_CF0703<\/p><div class=\"chromatogram-peaks\"><h4>Peaks<\/h4><table class=\"peaks col-lg-6 col-12 peak-50\">\n<thead><tr><th><\/th><th style=\"text-align: left;width: 75px\">Peaks<\/th><th style=\"text-align: center;width: 75px\">t<sub>R<\/sub> (min)<\/th><th style=\"text-align: center;width: 75px\">Precursor Ion<\/th><th style=\"text-align: center;width: 75px\">Product Ion<\/th><\/tr><\/thead>\n<tbody><tr><td class=\"num\">1.<\/td><td class=\"cmpd\">Norepinephrine-d6<\/td><td class=\"oth\">1.14<\/td><td class=\"oth\">158.16<\/td><td class=\"oth\">111.26<\/td><\/tr>\n<tr><td class=\"num\">2.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for Norepinephrine\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/51-41-2\/Norepinephrine\" target=\"_blank\" rel=\"noopener\">Norepinephrine<\/a><\/td><td class=\"oth\">1.14<\/td><td class=\"oth\">152.07<\/td><td class=\"oth\">106.91<\/td><\/tr>\n<tr><td class=\"num\">3.<\/td><td class=\"cmpd\">Epinephrine-d6<\/td><td class=\"oth\">1.32<\/td><td class=\"oth\">172.16<\/td><td class=\"oth\">111.86<\/td><\/tr>\n<tr><td class=\"num\">4.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for Epinephrine\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/51-43-4\/Epinephrine\" target=\"_blank\" rel=\"noopener\">Epinephrine<\/a><\/td><td class=\"oth\">1.33<\/td><td class=\"oth\">166.07<\/td><td class=\"oth\">135.00<\/td><\/tr>\n<tr><td class=\"num\">5.<\/td><td class=\"cmpd\">Normetanephrine-d3<\/td><td class=\"oth\">1.51<\/td><td class=\"oth\">169.00<\/td><td class=\"oth\">136.96<\/td><\/tr>\n<tr><td class=\"num\">6.<\/td><td class=\"cmpd\">Normetanephrine<\/td><td class=\"oth\">1.52<\/td><td class=\"oth\">166.00<\/td><td class=\"oth\">148.99<\/td><\/tr>\n<\/tbody><\/table>\n<table class=\"peaks col-lg-6 col-12 peak-50\"><thead><tr><th><\/th><th style=\"text-align: left;width: 75px\">Peaks<\/th><th style=\"text-align: center;width: 75px\">t<sub>R<\/sub> (min)<\/th><th style=\"text-align: center;width: 75px\">Precursor Ion<\/th><th style=\"text-align: center;width: 75px\">Product Ion<\/th><\/tr><\/thead><tbody>\n<tr><td class=\"num\">7.<\/td><td class=\"cmpd\">Dopamine-d4<\/td><td class=\"oth\">1.55<\/td><td class=\"oth\">141.16<\/td><td class=\"oth\">94.69<\/td><\/tr>\n<tr><td class=\"num\">8.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for Dopamine\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/51-61-6\/Dopamine\" target=\"_blank\" rel=\"noopener\">Dopamine<\/a><\/td><td class=\"oth\">1.56<\/td><td class=\"oth\">136.97<\/td><td class=\"oth\">64.66<\/td><\/tr>\n<tr><td class=\"num\">9.<\/td><td class=\"cmpd\">Metanephrine-d3<\/td><td class=\"oth\">1.94<\/td><td class=\"oth\">183.00<\/td><td class=\"oth\">151.15<\/td><\/tr>\n<tr><td class=\"num\">10.<\/td><td class=\"cmpd\"><a class=\"cmpd_link\" title=\"View compound information for Metanephrine\" href=\"https:\/\/ez.restek.com\/compound\/view\/en\/5001-33-2\/Metanephrine\" target=\"_blank\" rel=\"noopener\">Metanephrine<\/a><\/td><td class=\"oth\">1.96<\/td><td class=\"oth\">179.94<\/td><td class=\"oth\">165.01<\/td><\/tr>\n<tr><td class=\"num\">11.<\/td><td class=\"cmpd\">3-Methoxytyramine-d4<\/td><td class=\"oth\">2.42<\/td><td class=\"oth\">155.07<\/td><td class=\"oth\">122.93<\/td><\/tr>\n<tr><td class=\"num\">12.<\/td><td class=\"cmpd\">3-Methoxytyramine<\/td><td class=\"oth\">2.45<\/td><td class=\"oth\">151.00<\/td><td class=\"oth\">91.02<\/td><\/tr>\n<\/tbody><\/table><\/div><div class=\"chromatogram-conditions\"><h4>Conditions<\/h4><div class=\"conditions-container container-fluid\"><div class=\"row\"><table class=\"conditions col-lg-6 col-12\"><tr><th class=\"conditions_header\" scope=\"row\">Column<\/th><td>Raptor Biphenyl  (<a target=\"_blank\" href=\"https:\/\/www.restek.com\/p\/9309A62?utm_source=chromatograms&amp;utm_medium=link&amp;utm_campaign=LC_CF0703\" rel=\"noopener\">cat.# 9309A62<\/a>)<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Dimensions:<\/th><td>150 mm x 2.1 mm ID<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Particle Size:<\/th><td>2.7 \u00b5m<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Pore Size:<\/th><td>90 \u00c5<\/td><\/tr><tr><td><\/td><tr><th class=\"sub conditions_header\" scope=\"row\">Temp.:<\/th><td>30 \u00b0C<\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Standard\/Sample<\/th><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Conc.:<\/th><td>  Endogenous level<\/td><\/tr><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Inj. Vol.:<\/th><td>2 \u00b5L <\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Mobile Phase<\/th><td><\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">A:<\/th><td>Water, 0.2% formic acid <\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">B:<\/th><td>Methanol, 0.2% formic acid <\/td><\/tr><tr><td><\/td><td><table class=\"cgram_ramp\"><thead><tr><th>Time (min)<\/th><th>Flow (mL\/min)<\/th><th>%A<\/th><th>%B<\/th><\/tr><\/thead><tbody><tr><td>0.00<\/td><td>0.3<\/td><td>95<\/td><td>5<\/td><\/tr><tr><td>2.50<\/td><td>0.3<\/td><td>75<\/td><td>25<\/td><\/tr><tr><td>3.00<\/td><td>0.3<\/td><td>5<\/td><td>95<\/td><\/tr><tr><td>3.01<\/td><td>0.3<\/td><td>95<\/td><td>5<\/td><\/tr><tr><td>5.00<\/td><td>0.3<\/td><td>95<\/td><td>5<\/td><\/tr><\/tbody><\/table><\/td><\/tr><\/table><table class=\"conditions col-lg-6 col-12\"><tr><th class=\"conditions_header\" scope=\"row\">Detector<\/th><td>MS\/MS<\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Ion Mode:<\/th><td>ESI+ <\/td><\/tr><tr><th class=\"sub conditions_header\" scope=\"row\">Mode:<\/th><td>MRM <\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Instrument<\/th><td>UHPLC<\/td><\/tr><tr class=\"cgram_header_row\"><th class=\"conditions_header\" scope=\"row\">Sample Preparation<\/th><td>A 200 \u00b5L aliquot of urine sample (Bio-Rad Lyphochek quantitative urine control, normal level) was mixed with 10 \u00b5L (1 \u00b5g\/mL in methanol) internal standard solution and 600 \u03bcL of 250 mM ammonium acetate solution. The mixture was loaded onto the EVOLUTE EXPRESS WCX 96-well plate (30 mg), washed with 1 mL water and 1 mL methanol:acetonitrile (60:40), eluted with 200 \u00b5L of 5% formic acid in water:methanol (95:5), and injected (2 \u03bcL) for analysis.<\/td><\/tr><\/table><\/div><\/div><\/div><div class=\"chromatogram-pdf-link\"><a href=\"https:\/\/ez.restek.com\/images\/cgram\/lc_cf0703.pdf\" target=\"_blank\" rel=\"noopener noreferrer\"><svg xmlns=\"http:\/\/www.w3.org\/2000\/svg\" width=\"18\" height=\"18\" viewBox=\"0 0 18 18\"><g data-name=\"Group 2996\"><path data-name=\"Rectangle 1246\" d=\"M0 0h18v18H0z\" style=\"fill: none;\"><\/path><\/g><g data-name=\"Group 2997\"><path data-name=\"Path 729\" d=\"M13.412 11.4v2.017H5.345V11.4H4v2.017a1.349 1.349 0 0 0 1.345 1.345h8.068a1.349 1.349 0 0 0 1.345-1.345V11.4zm-.672-2.694-.948-.948-1.741 1.735V4H8.706v5.493L6.965 7.758l-.948.948 3.361 3.361z\" transform=\"translate(-.437 -.414)\" style=\"fill: rgb(13, 123, 196);\"><\/path><\/g><\/svg>Download PDF<\/a><\/div><\/div><\/div>\n<\/div><\/div><\/div>\n\n\n<p><\/p>\n\n\n\n<h3 class=\"wp-block-heading\">Linearity<\/h3>\n\n\n\n<p>Good linearity was achieved for all compounds using a 1\/x weighted linear regression for epinephrine, norepinephrine, dopamine, metanephrine, and normetanephrine, and a 1\/x quadratic regression for 3-methoxytyramine (Figure 3). All six analytes had r<sup>2<\/sup>&nbsp;values of 0.999 or greater and deviations of &lt;10%, except for the lowest concentration standard, which had deviations of &lt;20%.<\/p>\n\n\n<div class=\"wp-block-custom-chromatogram-article-top\"><div class=\"chromatogram-article-placeholder\"><div class=\"figure-heading\"><strong>Figure 3:<\/strong>\u00a0Calibration Curves.<\/div><div class='chromatogram-article-inner-full'><div class=\"chromatogram-article-inner\">\n<style>.kb-image43346_22989f-5e .kb-image-has-overlay:after{opacity:0.3;}<\/style>\n<div class=\"wp-block-kadence-image kb-image43346_22989f-5e\"><figure class=\"aligncenter size-large\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"876\" src=\"https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03-1024x876.jpg\" alt=\"\" class=\"kb-img wp-image-18910\" title=\"-\" srcset=\"https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03-1024x876.jpg 1024w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03-300x257.jpg 300w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03-768x657.jpg 768w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03-1536x1314.jpg 1536w, https:\/\/discover.restek.com\/wp-content\/uploads\/figure-article-CFAN2800-03.jpg 1800w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure><\/div>\n\n<\/div><\/div><\/div>\n\n\n<p><\/p>\n\n\n\n<h3 class=\"wp-block-heading\">Accuracy and Precision<\/h3>\n\n\n\n<p>Precision and accuracy analyses were performed on three different days. The accuracy of the method was demonstrated by the recovery values, which were within 10% of the nominal concentration for all QC levels. The %RSD was 0.2-6.2% and 1.8-3.8% for intraday and interday comparisons, respectively, indicating that method precision was also acceptable (Table II).<\/p>\n\n\n\n<p><strong>Table II:<\/strong>&nbsp;Accuracy and Precision of QC Samples<\/p>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><tbody><tr><td>&nbsp;<\/td><td colspan=\"3\"><strong>QC Level 1 (2.5\u201375 ng\/mL)<\/strong><\/td><td colspan=\"3\"><strong>QC Level 2 (25\u2013750 ng\/mL)<\/strong><\/td><td colspan=\"3\"><strong>QC Level 3 (75\u20131500 ng\/mL)<\/strong><\/td><\/tr><tr><td><strong>Analyte<\/strong><\/td><td><strong>Average Conc. (ng\/mL)<\/strong><\/td><td><strong>Average Accuracy (%)<\/strong><\/td><td><strong>%RSD<\/strong><\/td><td><strong>Average Conc. (ng\/mL)<\/strong><\/td><td><strong>Average Accuracy (%)<\/strong><\/td><td><strong>%RSD<\/strong><\/td><td><strong>Average Conc. (ng\/mL)<\/strong><\/td><td><strong>Average Accuracy (%)<\/strong><\/td><td><strong>%RSD<\/strong><\/td><\/tr><tr><td>Epinephrine<\/td><td>2.43<\/td><td>97.3<\/td><td>2.97<\/td><td>26.0<\/td><td>104<\/td><td>2.31<\/td><td>77.9<\/td><td>104<\/td><td>2.19<\/td><\/tr><tr><td>Norepinephrine<\/td><td>2.44<\/td><td>97.5<\/td><td>2.34<\/td><td>25.3<\/td><td>101<\/td><td>2.92<\/td><td>76.9<\/td><td>103<\/td><td>2.48<\/td><\/tr><tr><td>Dopamine<\/td><td>74.5<\/td><td>99.4<\/td><td>3.42<\/td><td>787<\/td><td>105<\/td><td>3.85<\/td><td>1,560<\/td><td>104<\/td><td>3.37<\/td><\/tr><tr><td>Metanephrine<\/td><td>25.4<\/td><td>102<\/td><td>2.46<\/td><td>81.0<\/td><td>108<\/td><td>1.81<\/td><td>774<\/td><td>103<\/td><td>2.00<\/td><\/tr><tr><td>Normetanephrine<\/td><td>78.4<\/td><td>105<\/td><td>3.36<\/td><td>788<\/td><td>105<\/td><td>2.98<\/td><td>1,570<\/td><td>105<\/td><td>3.52<\/td><\/tr><tr><td>3-Methoxytyramine<\/td><td>25.7<\/td><td>103<\/td><td>3.39<\/td><td>81.4<\/td><td>109<\/td><td>2.87<\/td><td>764<\/td><td>102<\/td><td>2.94<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<h3 class=\"wp-block-heading\">Urine Analysis<\/h3>\n\n\n\n<p>From three sets of analyses of blank and fortified urine samples (BioRad Lyphochek quantitative urine control, level 1 and level 2), the results showed that the recoveries of all fortified samples were within 90-105% of their nominal values (Table III). This demonstrated that the method is suitable for the analysis of catecholamines and metanephrines in urine from human patients at clinically relevant levels.<\/p>\n\n\n\n<p><strong>Table III:<\/strong>&nbsp;Analysis of Fortified Urine Samples<\/p>\n\n\n\n<figure class=\"wp-block-table\"><table class=\"has-fixed-layout\"><tbody><tr><td>&nbsp;<\/td><td colspan=\"5\"><strong>Level 1<\/strong><\/td><td colspan=\"5\"><strong>Level 2<\/strong><\/td><\/tr><tr><td><strong>Analyte<\/strong><\/td><td><strong>Blank Urine (ng\/mL)<\/strong><\/td><td><strong>Fortified Urine (ng\/mL)<\/strong><\/td><td><strong>Calculated Conc. (ng\/mL)<\/strong><\/td><td><strong>Fortified Conc. (ng\/mL)<\/strong><\/td><td><strong>Average Accuracy (%)<\/strong><\/td><td><strong>Blank Urine (ng\/mL)<\/strong><\/td><td><strong>Fortified Urine (ng\/mL)<\/strong><\/td><td><strong>Calculated Conc. (ng\/mL)<\/strong><\/td><td><strong>Fortified Conc. (ng\/mL)<\/strong><\/td><td><strong>Average Accuracy (%)<\/strong><\/td><\/tr><tr><td>Epinephrine<\/td><td>13.0<\/td><td>22.6<\/td><td>9.57<\/td><td>10.0<\/td><td>95.7<\/td><td>91.7<\/td><td>143<\/td><td>51.4<\/td><td>50.0<\/td><td>103<\/td><\/tr><tr><td>Norepinephrine<\/td><td>42.2<\/td><td>51.3<\/td><td>9.14<\/td><td>10.0<\/td><td>91.4<\/td><td>185<\/td><td>234<\/td><td>49.3<\/td><td>50.0<\/td><td>98.7<\/td><\/tr><tr><td>Dopamine<\/td><td>77.2<\/td><td>166<\/td><td>90.2<\/td><td>100<\/td><td>90.2<\/td><td>459<\/td><td>655<\/td><td>196<\/td><td>200<\/td><td>98.1<\/td><\/tr><tr><td>Metanephrine<\/td><td>71.9<\/td><td>163<\/td><td>90.7<\/td><td>100<\/td><td>90.7<\/td><td>516<\/td><td>699<\/td><td>183<\/td><td>200<\/td><td>91.7<\/td><\/tr><tr><td>Normetanephrine<\/td><td>251<\/td><td>345<\/td><td>94.8<\/td><td>100<\/td><td>94.8<\/td><td>1,310<\/td><td>1,500<\/td><td>188<\/td><td>200<\/td><td>94.3<\/td><\/tr><tr><td>3-Methoxytyramine<\/td><td>12.2<\/td><td>21.5<\/td><td>9.31<\/td><td>10.0<\/td><td>93.1<\/td><td>418<\/td><td>603<\/td><td>185<\/td><td>200<\/td><td>92.6<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<h2 class=\"wp-block-heading\">Conclusion<\/h2>\n\n\n\n<p>As demonstrated here, the Raptor Biphenyl column provides good retention and accurate results for the simultaneous analysis of catecholamines and metanephrines in urine. With a fast, simple sample preparation procedure and just five minutes of chromatographic analysis time, the established method is suitable for high-throughput labs performing analysis of pheochromocytoma and paraganglioma.<\/p>\n\n\n\n<div style=\"height:30px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n        <div class=\"cpb\">\n            <h3 class=\"cpb-heading\">Products Mentioned<\/h3>\n            <hr class=\"cpb-heading-underline\" \/>\n            <div class=\"cpb-list\">\n                                    <div class=\"cpb-item\">\n                        <div class=\"cpb-col cpb-col--left\">\n                            <a class=\"cpb-catalog\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/de.restek.com\/p\/9309A62\">                                Catalog No. 9309A62                            <\/a>                        <\/div>\n                        <div class=\"cpb-col cpb-col--middle\">\n                            <div class=\"cpb-title\">Raptor Biphenyl, 2.7 \u00b5m, 150&#215;2.1 mm HPLC-S\u00e4ule<\/div>\n                        <\/div>\n                        <div class=\"cpb-col cpb-col--right\">\n                                                            <a class=\"cpb-view-btn\" target=\"_blank\" rel=\"noopener noreferrer\" href=\"https:\/\/de.restek.com\/p\/9309A62\">Produkt anzeigen<\/a>\n                                                    <\/div>\n                    <\/div>\n                            <\/div>\n        <\/div>\n        \n\n\n<div style=\"height:100px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n","protected":false},"excerpt":{"rendered":"<p>Clinical diagnosis of pheochromocytoma and paraganglioma is often based on the analysis of catecholamines (epinephrine, norepinephrine, dopamine) and metanephrines (metanephrine, normetanephrine, 3-methoxytyramine) in urine. Analysis of these polar compounds using reversed-phase LC can be difficult due to limited chromatographic retention, which results in poor separation of the analytes from closely eluting matrix interferences. This method overcomes these problems by combining a simple solid phase extraction procedure with the consistent and accurate chromatographic performance of a Raptor Biphenyl column.<\/p>\n","protected":false},"author":46,"featured_media":6343,"comment_status":"closed","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"_acf_changed":false,"_kad_blocks_custom_css":"","_kad_blocks_head_custom_js":"","_kad_blocks_body_custom_js":"","_kad_blocks_footer_custom_js":"","_kadence_starter_templates_imported_post":false,"_kad_post_transparent":"","_kad_post_title":"","_kad_post_layout":"","_kad_post_sidebar_id":"","_kad_post_content_style":"","_kad_post_vertical_padding":"","_kad_post_feature":"","_kad_post_feature_position":"","_kad_post_header":false,"_kad_post_footer":false,"footnotes":""},"categories":[13],"tags":[],"industries-application":[2266,2267],"post-badge":[],"resource-type":[],"product-library":[2571,2553],"resource-technique":[2343,2365,2346],"ppma_author":[578,586],"class_list":["post-43346","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-application-notes","industries-application-clinical-ja","industries-application-clinical-diagnostics-ja","product-library-lc-columns-ja","product-library-liquid-chromatography-products-ja","resource-technique-liquid-chromatography-ja","resource-technique-ms-ms-ja","resource-technique-solid-phase-extraction-spe-ja"],"acf":[],"taxonomy_info":{"category":[{"value":13,"label":"Application 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