HPLC quantification of 5‐hydroxyeicosatetraenoic acid in human lung cancer tissues

Abstract

A simple and cost-effective HPLC method was established for quantification of 5-hydroxyeicosatetraenoic acid (5-HETE) in human lung cancer tissues. 5-HETE from 27 patients' lung cancer tissues were extracted by solid-phase extraction and analyzed on a Waters Symmetry C(18) column (4.6 x 250 mm, 5 microm) with a mobile phase consisting of methanol, 10 mM ammonium acetate, and 1 M acetic acid (70:30:0.1, v:v:v) at a flow rate of 1.0 mL/min. The UV detection wavelength was set at 240 nm. The calibration curve was linear within the concentration range from 10 to 1000 ng/mL (r(2) > 0.999, n = 7), the limit of detection was 1.0 ng/mL and the limit of quantitation was 10.0 ng/mL for a 100 microL injection. The relative error (%) for intra-day accuracy was from 93.14 to 112.50% and the RSD (%) for intra-day precision was from 0.21 to 2.60% over the concentration range 10-1000 ng/mL. By applying this method, amounts of 5-HETE were quantitated in human lung cancer tissues from 27 human subjects. The established HPLC method was validated to be a simple, reliable and cost-effective procedure that can be applied to conduct translational characterization of 5-HETE in human lung cancer tissues.

Role of LTB4in the pathogenesis of elastase-induced murine pulmonary emphysema

“…Lipid from the supernatant was extracted by C 18 Sep-pak cartridges (Waters) after preconditioning it with 100% HPLC methanol (JT Baker) three times and 100% HPLC H 2O (JT Baker) three times. After initially passing the lung homogenate samples through the cartridges, C18 cartridges were washed three times with 60% methanol-40% H2O solution, and then the lipid was eluted with 100% methanol-0.1% 1 N acetic acid (30). Eluted methanol was dried completely under vacuum in a Speedvac.…”

5‐lipoxygenase as an endogenous modulator of amyloid beta formation in vivo

“…The isocratic mobile phase consisted of methanol‐10mM ammonium acetate‐1M acetic acid (70:30:0.1; v:v:v), and the flow rate was 1.0ml/minute. The 5(S)‐HETE peak was identified and quantitated by ultraviolet (UV) absorbance at 240–270nm, respectively 13. LTB4, PGE2, and 12‐HETE were assayed by standardized enzyme immunoassays (EIAs), following the manufacturer's instructions (Assay Designs) and as described 4, 7, 14…”

Serum Free Fatty Acid Biomarkers of Lung Cancer

“…[22][23][24][25][26] Th e hydroxyeicosatetraenoic acids (HETEs) and hydroxyoctadecadienoic acids (HODEs) are stable oxidative products of arachidonic acid (AA) and LA, respectively. Th ere is considerable basic science evidence that FFAs and the oxidized FFAs HETE and HODE promote tumor development, progression, and metastases in lung cancer using cell culture, 27,28 tissues, [29][30][31] and animal models. 27,[32][33][34] For example, 5-HETE augments lung cancer cell survival, 30,35 12-HETE enhances lung cancer cell adhesion, [36][37][38][39] and 15-HETE promotes human lung cancer metastasis 40 and induces angiogenesis.…”

“…Th ere is considerable basic science evidence that FFAs and the oxidized FFAs HETE and HODE promote tumor development, progression, and metastases in lung cancer using cell culture, 27,28 tissues, [29][30][31] and animal models. 27,[32][33][34] For example, 5-HETE augments lung cancer cell survival, 30,35 12-HETE enhances lung cancer cell adhesion, [36][37][38][39] and 15-HETE promotes human lung cancer metastasis 40 and induces angiogenesis. 41,42 HODE is involved in many types of cancer, although its specifi c role remains unclear.…”