Abstract: Given the escalating global energy consumption, the inadequacy of resource reserves, and the increasingly prominent issue of environmental pollution, employing green materials for grease formulation holds immense significance in achieving sustainable development for both the economy and the environment. This work aimed to discover a new type of green thickener to broaden the application range of renewable and biodegradable materials in lubricating grease preparation. We had found that poly (3-hydroxybutyrate-co-4-hydroxybutyrate) (P34HB) possessed biodegradability, gel-forming ability and excellent mechanical properties by consulting literature. It was a promising petroleum-based material substitute in various fields. Moreover, in various fields of study, we had observed a remarkable similarity between the microstructure of the P34HB material employed for sample preparation and that of conventional lithium-based greases. For instance, electrospun P34HB exhibited a fibrous architecture. Collectively, we positted that the P34HB material exhibited potential for augmenting the viscosity of base oil. In this study, a range of novel greases with varying concentrations of P34HB as the thickener and castor oil as the base oil was prepared. The impacts of different thickener concentrations on physicochemical, rheological, and tribological properties were comprehensively investigated. The findings demonstrated that the colloid stability and oxidation stability of P34HB lubricating grease improved with an increase in thickening agent content. The SEM analysis revealed that the microstructure of the P34HB grease thickener differed from the fiber helical structure observed in lithium-based grease, exhibiting an irregular stacking sheet structure akin to bentonite and certain polyurea greases. The formation of this form might be attributed to the hydrogen bonding between P34HB and castor oil, as evidenced by the Fourier infrared spectrum and Raman spectral analysis. This interaction leaded to irregular accumulation and aggregation of sheet materials, thereby impeding the flow of base oil. The rheological property test results additionally demonstrated that the P34HB grease system exhibited shear-thinning behavior and adhered to the Carreau Yasuda model (R²>0.999), thereby confirming its non-Newtonian properties akin to conventional greases available in the market. Additionally, the friction behavior results demonstrated that under mild friction conditions, the P34HB lubricating grease with a thickening agent content of 48% demonstrated frictional properties comparable to the control group, castor oil lithium-based grease. The preliminary analysis results of the lubrication mechanism of P34HB, combined with SEM and XPS techniques, revealled that the thickener microstructure in the P34HB grease system exhibited a relatively thinner sheet-like morphology (with a thickness less than 100 nm). Consequently, it could be inferred that during the friction process, a significant number of small-sized sheet structures were continuously transferred to the surface of the friction pair through the base oil under frictional forces. This transfer effectively supported and maintained an adequate thickness of the oil film within the contact area, working synergistically with subsequent chemical reaction films to provide lubrication. This study provided an effective strategy for the application of P34HB in the lubrication field and paved the way for the development of a new generation of environmentally friendly lubricating greases.