Following the signing of the Tobacco 21 Amendment (T21) in December 2019 to raise the minimum legal age for the sale of tobacco products from 18 to 21 years in the United States, there is a need to monitor public responses and potential unintended consequences. Social media platforms, such as Twitter (subsequently rebranded as X), can provide rich data on public perceptions. This study contributes to the literature using Twitter data to assess the knowledge and beliefs of T21. Twitter data were collected from November 2019 to February 2021 using the Twitter streaming application programming interface with keywords related to vaping or e-cigarettes, such as "vape," "ecig," etc. The temporal trend of the T21 discussion on Twitter was examined using the mean number of daily T21-related tweets. Inductive methods were used to manually code the tweets into different sentiment groups (positive, neutral, and negative) based on the attitude expressed toward the policy by 3 coders with high interrater reliability. Topics discussed were examined within each sentiment group through theme analyses. Among the collected 3197 tweets, 2169 tweets were related to T21, of which 444 tweets (20.5%) showed a positive attitude, 736 (33.9%) showed a negative attitude, and 989 (45.6%) showed a neutral attitude. The temporal trend showed a clear peak in the number of tweets around January 2020, following the enactment of this legislation. For positive tweets, the most frequent topics were "avoidance of further regulation" (120/444, 27%), "Enforce T21" (110/444, 24.8%), and "health benefits" (81/444, 18.2%). For negative tweets, the most frequent topics were "general disagreement or frustration" (207/736, 28.1%) and "will still use tobacco" (188/736, 25.5%). Neutral tweets were primarily "public service announcements (PSA) or news posts" (782/989, 79.1%). Overall, we find that one-third of tweets displayed a negative attitude toward T21 during the study period. Many were frustrated with T21 and reported that underage consumers could still obtain products. Social media data provide a timely opportunity to monitor public perceptions and responses to regulatory actions. Continued monitoring can inform enforcement efforts and potential unintended consequences of T21.