Early one morning, Fern Arable (Pamelyn Ferdin) prevents her father (John Stephenson) from slaughtering a piglet as therunt of the litter. Deciding to let her daughter deal with nurturing a pig, John Arable spares the piglet, and allows her daughter to raise it as a pet. Fern nurtures it lovingly, naming it Wilbur. Six weeks later, Wilbur, due to being a spring pig, has matured, and John decides to sell Wilbur to Fern's uncle, Homer Zuckerman (Bob Holt). At the farm, Wilbur (Henry Gibson) speaks his first words, though he is left yearning for companionship, and attempts to get Templeton (Paul Lynde) and the goose (Agnes Moorehead) to play with him, though they refuse. Wilbur then wants a lamb to play with him, though the lamb's father (Dave Madden) says sheep don't play with pigs because it's only a matter of time before he will be slaughtered, and turned into smoked bacon and ham. Wilbur reduces himself to tears until a mysterious voice tells him to "chin up", and wait until the morning to reveal herself to him. The next morning, the voice sings a song about "chinning up", and reveals herself to be a Araneus cavaticus named Charlotte (Debbie Reynolds), living on a web overlooking Wilbur's enclosure. Charlotte tells Wilbur that she will come with a plan guaranteed to spare his life.
After Wilbur makes friend with a young gosling named Jeffrey (Don Messick), Charlotte reveals her plan to "pay a trick on Zuckerman", and consoles Wilbur to sleep. The next morning, Zuckerman's farm assistant sees the words, SOME PIG, spun within Charlotte's web. The incident attracts publicity among Zuckerman's neighbors who attribute the praise to a divine intervention. The publicity eventually dies down, and Charlotte requests the barn animals to devise a new word to spin within her cobweb. After several suggestions, the goose suggests the phrase, TERRIFIC! TERRIFIC! TERRIFIC!, though Charlotte decides to shorten it to one TERRIFIC. The incident becomes another media sensation, though Zuckerman still desires to slaughter Wilbur. For the next message, Charlotte then employs Templeton to pull a word from a magazine clipping for inspiration, in which Templeton returns the word, RADIANT, ripped from a soap box to spin within her cobweb. Following this, Zuckerman decides to enter Wilbur into the county fair for the summer. Charlotte decides to accompany Wilbur, though Templeton is reluctant until he is convinced by the goose about the food at the fair. After one night at the fair, Charlotte sends Templeton on another errand to gather another word for her next message, in which Templeton returns with the word, HUMBLE. The next morning, Wilbur awakens to find Charlotte has spun an egg sac containing her unborn offspring, and the following afternoon, the word, HUMBLE, is spun. However, Avery (Danny Bonaduce) discovers another pig named Uncle has won first place, though the county fair staff decides to hold a celebration in honor of Zuckerman's miraculous pig, and rewards Zuckerman $25 and a gold metal. Zuckerman then announces that he will allow Wilbur to live.
Exhausted from laying eggs and messages, Charlotte tells Wilbur she will remain at the fair to die. Following her soothing song of "Mother Earth and Father Time", Wilbur has Templeton to retrieve Charlotte's egg sac to take back to the farm, and Charlotte dies soon after. Having returned to Zuckerman's farm, Wilbur guards Charlotte's egg sac until the winter. The next spring, Charlotte's 5,000 children are hatched, and leave the farm causing Wilbur to become saddened though three of Charlotte's daughters stay behind. Pleased at finding new friends, Wilbur names the spiderlings Joy, Nellie, and Aranea, and the story concludes mentioning that more generations of spiders kept him company in subsequent years.
- Zecora, Stephen, Sharky, Mako, Willy, T.C., Hugs, Buzz, J.J., Socky, Skunky, 1206, The Miner Trains: (Steam Grindor, Steam Mech, Mucker, Steam Claw D., Steam Driller, Steam Sweeper, and Steam Exavator), Evan, Steamy, Puffy, Shai-Shay, Rattlesnake Jake, The Planet Trains: (Zip, Dazzlen, Dusten, Gustis, Rings, Uray, Tune, and Shiver), Dusty Crophopper, Bill and Ben, Rodney Copperbottom, Fender Pinwheeler, Wonderbot, Piper Pinwheeler, Crank Casey, Lug and Diesel guest star in this film.
- The storyline continues in Thomas and Twilight Sparkle's Adventures of Charlotte's Web 2: Wilbur's Great Adventure.
- Opening/life on a harm/Fern saves the runt
- "There Must Be Something More"/Henry Fussy
- Wilbur grows up
- Life at the Zuckermann's/"I Can Talk"
- Wilbur meets our heroes/Discovering Wilbur's dark future
- "Chin Up"/Meeting Charlotte
- The eggs hatch!/"We Got Lost in Common"
- Henry tries to catch Charlotte/"Deep in the Dark"
- "Some Pig"/news spreads
- New word/"Terrific"
- Searching for a new word
- "Radiant"/"Mother Earth Father Time"
- Preparing for the Fair/"A Veritable Smorgasboard"
- Getting Wilbur into the crate/Arriving at the Fair
- "There Must Be Something More" Reprise/Uncle the Pig
- Fun at the fair/A new word/"A Veritable Smorgasboard" Reprise
- The Next morning/Charlotte's egg sack/Templeton returns
- "Humble"/At the stand/"Zuckermann's Famous Pig"
- Charlotte passes
- Returning to the farm/Winter
- Spring time/Birth of Charlotte's daughters/Ending
- I Can Talk!
- Chin Up
- We've got Lots in Common
- Deep In The Dark/Charlotte's Web
- Mother Earth and Father Time
- A Veritable Smorgasbord
- Zuckerman's Famous Pig
- Pokémon: The First Movie - That Which Has Life (when everyone mourns over Charlotte)
- Ordinary Miracle (end credits song)