Valve-through tire puncture sealant compositions including dissolved and undissolved natural bio-materials, anti-freezing agents and other rigid particles are invented. In the preparation, an aqueous solution with 0.1-10 wt % dissolved bio-materials and 0.1%-10 wt % tackifier is mixed with anti-freezing agent, in which 0.01-5 wt % rigid particles of bio-materials and other particles are suspended. Weight percentage of water and anti-freezing agent are 20-40 wt % and 20-70 wt % respectively. Trace amounts of additives such as coloring and preservative are possibly added to the sealant. The invention includes the method for preparing the sealant composition containing bio-materials. The proposed sealant compositions have several advantages: Environmentally friendly, insignificant settling, long shelf-life, etc. The sealant is capable of sealing puncture formed by less than or equal to 6.35 mm spike for long duration, i.e. more than 30 hours.