Abstract: Objective: To explore the factors affecting the content of gastrodin and p-hydroxybenzyl alcohol in fresh Gastrodia elata. Methods: This study optimized the pre-treatment sample preparation method and chromatographic conditions for the determination of gastrodin and p-hydroxybenzyl alcohol in fresh Gastrodia elata. The sample was homogenized, mixed with 87% ethanol, ultrasonic extraction, centrifugation, nitrogen blowing and water bath concentration. After passing through a 0.45 μm filter membrane, the sample was eluted equivalently on a C₁₈ column(4.6 mm×250 mm, 5 μm) with acetonitrile-0.05% phosphoric acid aqueous solution(3:97, V/V) as the mobile phase at 220 nm. The flow rate was 1.0 mL·min^-1 and the column temperature was 25℃, high performance liquid chromatography(HPLC) was used to analyze and determine the contents of gastrodin and p-hydroxybenzyl alcohol in 72 fresh Gastrodia elata collected from different size hierarchy, origins and cultivation methods in Huichuan district, Zunyi city. Results: The results of methodology validation showed that the linear relationship of gastrodin and p-hydroxybenzyl alcohol was good in the corresponding range under three spiked concentrations, R²>0.9999, the recovery was 81.20%~107.30%, RSD<5%. The total content of gastrodin and p-hydroxybenzyl alcohol in 72 samples was 0.262%~1.150%.The sample preparation method of fresh gastrodia elata used in this study is simple, rapid and has good extraction effect. The analytical method has good precision, reproducibility and separation effect, which is suitable for the study and analysis of gastrodin and p-hydroxybenzyl alcohol in fresh gastrodia elata. Through further analysis of the effects of different grades, sizes, origins, and different cultivation methods on the content of gastrodin and p-hydroxybenzyl alcohol in fresh gastrodia, the results showed that the higher the quality grade of fresh gastrodia, the higher the total content of gastrodin and p-hydroxybenzyl alcohol;Gastrodia planting bases are located at different altitudes, environmental vegetation types, geographic locations, and sunlight, and the development time of Gastrodia planting under forests in each planting base is different. Therefore, the gastrodin and p-hydroxy There are also significant differences in the content of benzyl alcohol;and a comparative analysis of fresh Gastrodia elata with different cultivation methods reveals that sexual reproduction is better than asexual reproduction. Conclusion: The influence of gastrodin and p-hydroxybenzyl alcohol in fresh Gastrodia elata. in different origins, cultivation methods and size hierarchy in Huichuan district has certain differences.