Leaves preparations of Persea americana Mill., Lauraceae, popularly known as abacateiro, have been used as antidiabetic, hypolipidemic and hypotensive1,2. However, the quality control parameters for its fluid extract uses old fashionable techniques and needs improvement. In this way, the objective of this study was to determine quality control parameters by HPLC and to optimize hydrolysis conditions for quantification of flavonoids expressed in quercetin in the fluid extract of P. americana. Six samples of fluid extracts were prepared by percolation method with ethanol 70% (v/v) and one sample was obtained commercially. The relative density, dry residue, chromatographic conditions for thin-layer chromatography and high-performance liquid chromatography were determined. In addition, the hydrolysis conditions were optimized for the quantification of flavonoid content in the fluid extract. Among the quality control parameters, the relative density of the fluid extracts varied from 0.95231 to 1.10382 and the dry residue varied from 13.4449 to 26.2505 % (w/w) (RSD maximum: 5%). Chromatographic conditions for thin layer chromatography were: stationary phase: silica gel GF254 (200 μm), mobile phase: ethyl acetate, isopropyl alcohol, water and formic acid (300: 17: 13: 0.1), sample: 10 μl of fluid extract (conc. 100 μl/ml), standard: 10 μl of mixture of quercetin (0.25 mg/ml) and rutin (1mg/ml) diluted in methanol and detection with 2-aminoethyldiphenylborinate (1% in ethanol), followed by polyethylene glycol 4000 (5% in methanol) and UV light (365 nm). For high performance liquid chromatography with UV/Vis detector (371 nm), the parameters were standardized: sample (conc. 14.28 μl/ml in methanol: water 1:1), standard: quercetin (conc. 0.038 mg/ml in methanol), injection volume = 20 μl, stationary phase: C18 column (250 mm x 4.6 mm x 5 μm), pre-column C18, 5 μm, temperature at 20 °C, flow 1 ml/min, with mobile phase A: water acidified with phosphoric acid (0.1%) and phase B: acetonitrile, using the following conditions: phase A = 0 ' -> 20' = 55% -> 5%; 20 ' -> 25' = 5% -> 5%, with an equilibrium time of 10 min. Due the occurrence of different glycosylated flavonoids, the flavonoid content was performed after the hydrolysis to quantify flavonoids expressed in quercetin. In order to optimize the hydrolysis conditions was performed factorial design 23 with three factors tested in two levels of variation (-1 and +1), with a central point (0) in duplicate, varying the hydrochloric acid volume (0.5 ml to 1.0 ml), hydrolysis time (30 to 60 min), methanol concentration (50 to 100), considering as response the quercetin concentration, analyzed by external calibration (y = 8 108 x – 343618; r2: 0.9983). The t-test (95% confidence) showed that were significant the acid variable (where higher acid volume, higher content of quercetin) and the acid-x solvent interaction. The highest content was obtained with 1 ml of concentrated hydrochloric acid and 50% methanol. When the acid concentration was increased the content was increased. The hydrolysis conditions were defined as: 500 μl of fluid extract, 1.5 ml of concentrated hydrochloric acid and 20 ml of methanol 75% under reflux at 80-90 °C for 30 min. The contents varied from 0.01702 to 0.1517 g of quercetin per 100 g of sample. Considering the importance in the development of quality control parameters for the commercialization of products, this study developed pharmacopoeial parameters aiming to contribute to the guarantee of standardization and quality.