The main characters of the novel are Florentino Ariza and Fermina Daza. Both Florentino and Fermina fall in love with each other in their youth. A secret relationship blossoms between the two with the help of Fermina's Aunt Escolástica. They exchange several love letters. However, once Fermina's father, Lorenzo Daza, finds out about the two, he forces his daughter to stop seeing Florentino immediately. When she refuses, he and his daughter move in with his deceased wife's family in another city. Regardless of the distance, Fermina and Florentino continue to communicate via telegraph. However, upon her return, Fermina realizes that her relationship with Florentino was nothing but a dream since they are practically strangers; she breaks off her engagement to Florentino and returns all his letters.
A young and accomplished national hero, Dr. Juvenal Urbino, meets Fermina and begins to court her. Despite her initial dislike of Urbino, Fermina gives in to her father's persuasion and the security and wealth marrying Urbino offers, and they wed. Urbino is a medical doctor devoted to science, modernity, and "order and progress". He is committed to the eradication of cholera and to the promotion of public works. He is a rational man whose life is organized precisely and who greatly values his importance and reputation in society. He is a herald of progress and modernization.
Even after their engagement and marriage, Florentino swore to stay faithful and wait for Fermina. However, his promiscuity gets the better of him. Even with all the women he is with, he makes sure that Fermina will never find out. Meanwhile, Fermina and Urbino grow old together, going through happy years and unhappy ones and experiencing all the reality of marriage. At an elderly age, Urbino attempts to get his pet parrot out of his mango tree, only to fall off the ladder he was standing on and die. After the funeral, Florentino proclaims his love for Fermina once again and tells her he has stayed faithful to her all these years. Hesitant at first because of the advancements he made to a newly made widow, Fermina eventually gives him a second chance. They attempt a life together, having lived two lives separately for over five decades.
Urbino's function in the novel is to contrast with Florentino Ariza and his archaic and boldly romantic love. Urbino proves in the end not to have been an entirely faithful husband, confessing one affair to Fermina many years into their marriage. Though the novel seems to suggest that Urbino's love for Fermina was never as spiritually chaste as Florentino Ariza's was, it also complicates Florentino's devotion by cataloging his many trysts as well as a few potentially genuine loves. By the end of the book, Fermina comes to recognize Florentino's wisdom and maturity and their love is allowed to blossom during their old age.
- Lorenzo Daza – Fermina Daza's father, a mule driver; he despised Florentino and forced him to stop meeting Fermina. He is revealed to have been involved in some illicit businesses to build his fortune.
- Jeremiah de Saint-Amour – The man whose suicide is introduced as the opening to the novel; a photographer and chess-player.
- Aunt Escolástica – The woman who attempts to aid Fermina in her early romance with Florentino by delivering their letters for them. She is ultimately sent away by Lorenzo Daza for this.
- Tránsito Ariza – Florentino's mother.
- Hildebranda Sánchez – Fermina's cousin.
- Miss Barbara Lynch – The woman with whom Urbino confesses having had an affair, the only one during his long marriage.
- Leona Cassiani – She starts out as the "personal assistant" to Uncle Leo XII at the R.C.C., the company which Florentino eventually controls. At one point, it is revealed that the two share a deep respect, possibly even love, for each other, but will never actually be together. She has a maternal love for him as a result of his "charity" in rescuing her from the streets and giving her a job.
- Diego Samaritano – The captain of the riverboat on which Fermina and Florentino ride at the end of the novel.
- América Vicuña – The fourteen-year-old girl who towards the end of the novel is sent to live with Florentino; he is her guardian while she is in school. They have a sexual relationship, and after being rejected by Florentino and failing her exams, she kills herself. Her suicide illustrates the selfish nature of Florentino's love for Fermina.