## 4 Discussion and Conclusions  
Bio-materials, allowing for the use of renewable resources, are increasingly applied in industry due to sustainability policy. Once used, they cannot detrimentally change properties of the material, instead they should make it more ecologically friendly. The effect of initial fluidization of bitumen with the bio-agent and then the gradual polymerization suggests the potential of its application as a bi-functional additive in warm mix asphalt (WMA) technology or asphalt mixes with reclaimed asphalt pavement (RAP). Asphalt mixture production is a sensitive process requiring high quality control. During application of RAP directly to the mixer there is a potential risk related to the mixture uniformity. Application of bio-agent with fluxing function lowers bitumen viscosity and helps to reach better homogeneity in the mixtures containing RAP. It also should be highlighted that, in asphalt mixtures containing RAP there are two types of bitumen binder: binder from RAP and virgin bitumen. It is commonly assumed that during asphalt mixture production both bitumens will mix resulting with blended bitumen. The tests conducted shows that even low rate of bio-agent (2.5%) beneficially changes bitumen properties in low temperature; moreover polymerization processes occurring in second stage of the process improves bitumen properties in high operational temperature. The testing of complex modulus \(|\)G*\(|\) and phase angle \(|\)8\(|\) demonstrated that it is possible to select optimal bio-agent content depending on the desired consistency of bitumen. As mentioned earlier, bio-agent polymerization occurring during the second stage of the process partially recovers initial properties of the bitumen. Unrecovered parts of the fluxing effect compensate, in the blended RAP + virgin bitumen, the stiffening of the binder caused by the application of aged bitumen from RAP. Due to the fact that bio-additive modifies visco-elastic properties of bitumen in a wide temperature range and has a fluxing effect, it also improves the compaction process of the asphalt mixture. As shown in this research, the bio-agent influences the amount of air voids in the compacted mixture. Based on the observed compaction of mixtures with RAP as well as with RAP and bio-agent, it can be concluded that the design process of such mixes should include an expected effect of fluxing in order to obtain the desired air void content and compaction level. The fluxing effect of bitumen binder and the changes in its viscoelastic properties permanently influence the behavior of asphalt mixtures. Initially, before bio-agent polymerization, improvement in compactibility can be seen. Next, after polymerization, the mixture exhibits higher fatigue durability, accommodates higher strains, and shows improved low-temperature properties. Those effects are most likely caused by an improvement in both virgin binder properties and aged (RAP origin) binder properties. Asphalt mixtures with bio-agents added in the amount of 2.5% shows 10% higher fatigue durability (4 PB) and a 5 \({}^{\circ}\)C lower failure temperature (TSRST). As was demonstrated in another paper [15] (more chemically oriented) produced based on this study, the polymerization process of the bio-agent occurs until the saturation of double unsaturated  
Figure 8: Low temperature characteristic of asphalt mixtures: (**a**) failure temperature; (**b**) cryogenic stress. bonds, whose number is limited--typically, bio-agent polymerization occurs during the first month (with a complete stop after two months). For this reason, over-stiffening of the mixture during pavement in-service life is not expected. The bio-agent used in this research is a material obtained from RME with an average polymerization potential due to the limited number of unsaturated bonds.