The peptide linker between variable domains of heavy (V_H) and light (V_L) chains is one of important factors that influence the characteristics of scFv, including binding activity and specificity against target antigen. The scFvs against daidzin (DZ-scFvs) with different linker lengths were constructed in the format of V_H-(GGGGS)_n-V_L (n = 1, 3, 5, and 7). They were expressed in the hemolymph of silkworm larvae using the Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid DNA system, and their reactivity against daidzin and related compounds were evaluated using an indirect competitive enzyme-linked immunosorbent assay (icELISA), which is applicable for quantitative analysis of daidzin. The results showed that the reactivity of scFvs against daidzin was increased, whereas specificity slightly decreased when their peptide linker was lengthened. These results suggested that the linker length of DZ-scFvs contributes to its reactivity. In addition, the results emphasize that the linker length could control the reactivity of DZ-scFvs.

This study outlines a green process for Centella asiatica (L.) Urb. (CA) extraction. Natural deep eutectic solvents (NADESs) and microwave-assisted extraction (MAE) were combined to provide a high bioactive compound yield and high antioxidant activity. Among the NADESs evaluated, the combination of acetylcholine chloride : malic acid : water (1 : 2 : 2): water (40 : 60) was the best for extraction. These conditions provide high madecassoside (MS) (21.7 mg g⁻¹ dry weight) and asiaticoside (AS) (12.7 mg g⁻¹ dry weight) yields, with greater than 80% (v/v) EtOH (13.3 mg g⁻¹ MS and 7.80 mg g⁻¹ AS). In addition, the extracts from this process showed higher antioxidant activity (IC₅₀ = 0.26 mg mL⁻¹) than the CA aqueous EtOH and water extracts. Moreover, the color of the extract products was less green than that of the extracts prepared using EtOH and aqueous EtOH as solvents, which are suitable for cosmeceutical products. Response surface methodology (RSM) was used for MAE optimization. The ANOVA data from the central composition design (CCD) of RSM were fitted with quadratic models yielding acceptable R² (>0.93), adjusted R² (>0.87), predicted R² (>0.81), and nonsignificant lack of fit (p > 0.05) values. The quadratic model was validated using optimal conditions (30 s, power 300 W, and a liquid to solid ratio 20 mL g⁻¹), and the model validation showed more than 80% accuracy in both MS and AS yields. This research presented an effective green process for CA extraction, which resulted in an environmentally friendly CA extract requiring little energy consumption and no organic solvents.

Menthol and oleic acid are therapeutic hydrophobic deep eutectic solvents for solubilizing, delivering, and improving anti-inflammation properties of curcuminoids and curcumin with less cytotoxicity.