The protagonist of the game sees a strange dream: a black jaguar appears and introduces himself as a servant of the Inca king. He explains that the Inca people are dying of malaria and endangered by the European explorers. The jaguar is willing to transfer the hero to the past, where he/she will have to look for the cinchona plant and deliver it to the Inca king.

Part of the Trail series of educational games, The Amazon Trail features a gameplay system similar to the other installments, with adventure and simulation elements. The player will have to choose travel guides, make travel plans, manage supplies, navigate a boat on the Amazon river, encounter many indigenous animal species, take pictures, and catch fish in order to survive the hazards of the journey. At certain points in the game, time-traveling is also available, and the player is able to meet historical characters. Upon the completion of the main quest, players are graded according to their gameplay style, taking into account personal activities, number of animals identified, additional gifts brought to the Inca king, etc.