Background: Today, learning English is an undeniable necessity because English is the language of science and the training of different disciplines at higher levels and science production is done in English. Motivation is a variable that is considered as a fundamental pillar in the learning category. Methods: In this cross-sectional study, 607 students of Mashhad University of Medical Sciences and Ferdowsi University from faculties of Health, Nursing and midwifery, paramedical, medicine, dentistry, pharmacy, agriculture, geological, veterinary, and biology who were studying in the third semester were studied during the academic year of 2018-2019. In this study, the faculties of Mashhad University of Medical Sciences and Ferdowsi University were selected using a stratified sampling method. In addition, the motivation and attitude of the English language Learning (Gardner) questionnaire was used in the present study. Results: The mean score of total attitude and knowledge in Ferdowsi University was 109. 29 ± 16. 04, and in the University of Medical Sciences it showed that 108. 91 ± 17. 39. 2. 1% of the students had poor motivation and attitude, while it was medium in 24. 7%, good in 64. 7% of students, and 8. 4% had a very good motivation and attitude to learn. By assessing the scores of attitude and motivation, it was demonstrated that the students had a positive attitude towards the language; however, the motivation of language was not favorable among students. There was a significant relationship between students' motivation and their educational level (P < 0. 05); however, the students with higher educational levels were more motivated. There was no significant difference between students’ motivations of two universities. No significant relationship could be observed between motivation and major, native status, age, as well as gender (P > 0. 05). Conclusion: The students of two universities, without significant differences, had a good attitude toward language learning. However, the motivation score was not as desirable. Not using the main books and resources of language, inappropriate employment or lack of employment, and economic difficulties were considered as motivational barriers.