This study aimed to investigate the potential mitigation of copper stress in tomato plants (Solanum pimpinellifolium L0566) through the addition of chitosan (CH) or nanosilver (nAg) to Murashige and Skoog (MS) medium under in vitro conditions. Various growth parameters; the proline, malondialdehyde (MDA), total polyphenol and mineral contents; and CIE L*a*b* colour parameters were evaluated. After the multiplication stage, the explants were transferred to MS medium (control); MS + 20 ppm chitosan with a molecular weight of 3.33 kDa (CH3.33); MS + 6 mg l–1 nAg; MS + 100 μM l–1 copper sulfate (CuSO4); MS+20 ppm CH3.33+100 μM l–1 CuSO4; or MS + 6 mg l–1 nAg + 100 μM l–1 CuSO4. The results indicated that while CuSO4 or CuSO4-nAg solutions inhibited growth traits, CH3.33 stimulated growth, particularly shoot production, and plants treated with CH3.33 exhibited better developed roots and a higher fresh mass. Additionally, CH3.33 alleviated the negative effects of CuSO4 on the proline, MDA and total polyphenol contents in tomato plants. Moreover, tomato explants exhibited greener leaves, while those treated with nAg and CuSO4 showed decreased colour values. CH3.33 or nAg positively influenced the mineral content of tomato leaves under heavy metal stress. This study underscores the complex interactions between growth medium components on tomato plant growth, physiology and the mineral content, highlighting the potential of chitosan in mitigating heavy metal stress in tomato plants.