To develop an alternative high-protein forage resource to alleviate ruminant feed shortages, we investigated the effects of replacing alfalfa (Medicago sativa L.) with different ratios of paper mulberry (Broussonetia papyrifera L., RY) on fermentation quality, protein degradation, and in vitro digestibility of total mixed ration (TMR) silage. The TMR were made with alfalfa and RY mixtures (36.0%), maize meal (35.0%), oat grass (10.0%), soybean meal (7.5%), brewers’ grain (5.0%), wheat bran (5.0%), premix (1.0%), and salt (0.5%) on a dry matter basis, respectively. The alfalfa and RY mixtures were made in the following ratios of dry matter: 36:0 (RY0), 27:9 (RY9), 18:18 (RY18), 9:27 (RY27), and 0:36 (RY36). After ensiling for 7, 14, 28, and 56 days, fermentation quality, protein degradation, and microbial counts were examined, and chemical composition and in vitro digestibility were analyzed after 56 days of ensiling. All TMR silages, irrespective of the substitution level of RY, were well preserved with low pH and ammonia nitrogen content, high lactic acid content, and undetectable butyric acid. After ensiling, the condensed tannin content for RY18 silages was higher than the control, but non-protein nitrogen, peptide nitrogen, and free amino acid nitrogen contents was lower, while the fraction B1 (buffer-soluble protein) was not different among all the silages. Dry matter and crude protein digestibility for RY27 and RY36 silages was lower than the control, but there was no difference between control and RY18 silages. This study suggested that ensiling RY with alfalfa inhibited true protein degradation, but decreased in vitro dry matter and crude protein digestibility of TMR silages, and that 18:18 is the optimal ratio.

Humic acid (HA), as an important by-product, has been demonstrated to affect anaerobic digestion performance and subsequent land application of digestate via the batch anaerobic digestion process. However, the knowledge about the evolution of structure and function of HA during continuous anaerobic digestion (AD) is still unclear. Therefore, the current study examined the structural changes in HA produced during the continuous AD process and its metal-adsorption-reduction abilities. The results of three-dimensional fluorescence spectroscopy showed a general upsurge in humic-like components’ abundance (70–77%), with an increase in humification index (2.56–3.43). Likewise, the content of HA increased from 4.8 g L⁻¹ to 6.9 g L⁻¹ in the continuous AD process. The evolution of C-H, O-H, C=O, C=C, and C-O functional groups of HA was observed via the 2D COS FTIR analysis. Moreover, the concurrent dynamics of functional groups contributed to the higher adsorption (255.2 mg g⁻¹) of Cr (VI) and reduction (60.3 mg g⁻¹) of Cr (VI) to Cr (III) after 168 days of the continuous AD process. The findings of the current study not only advanced understanding of the evolution of HA during continuous anaerobic digestion and its metal remediation potential but also support further research toward developing an eco-friendly and innovative strategy for the remediation of heavy metals contaminated soils employing anaerobic digestate as an auxiliary agent.

A novel struvite–humic acid composite (S–HA) was derived from an anaerobic digestate and evaluated for the adsorption and reduction of chromium [Cr (VI)] in this study. The results indicated that the struvite–humic acid composite (S–HA) contains higher contents of oxygen-containing and aromatic functional groups (47.05% and 34.13%, respectively) and a higher specific surface area (19.3 m² g⁻¹). These special characteristics of S–HA contributed to its higher adsorption capacity (207.69 mg g⁻¹ and 254.47 mg g⁻¹ for pseudo-first and second-order kinetic models, respectively) for chromium. Furthermore, XPS analysis showed that a portion of the bonded Cr (VI) was reduced to Cr (III) by carboxyl and hydroxyl functional groups, which oxidized and changed into ketone and phenol functional groups. Based on the findings, it was concluded that the phosphate–humic acid composite has an outstanding chromium adsorptive and reduction capacity. However, more research is needed to fully understand the potential of the struvite–humic acid composite for chromium adsorption and reduction.

This study investigated the impact of different strains of lactic acid bacteria (LAB) on the vitamin A content in total mixed ration (TMR) silage. The TMRs were inoculated with Lactiplantibacillus plantarum (LP), Lacticaseibacillus casei (LC), Enterococcus faecium (EF) or left without LAB as control (CK) and subsequently ensiled under dark, anaerobic conditions for 56 days. Throughout the ensiling process, the fermentation quality, chemical composition, microbial community, and vitamin A content of the TMR silage were monitored. Additionally, the free radical scavenging ability, total antioxidant capacity (T-AOC), and activities of antioxidant enzymes (SOD, GSH-Px, CAT) in the three LAB strains were assessed. After 56 days of ensiling, the antioxidant enzyme activities of T-AOC, SOD, GSH-Px and CAT of TMR silage were also analyzed. The results indicated that LAB inoculation had a significant effect on both the fermentation quality and vitamin A content of TMR silage (p < 0.05). Among the LAB strains, LP and LC enhanced the fermentation quality of the TMR silage. Vitamin A loss rates in decreasing order were LC, CK, EF and LP groups. Notably, LAB strains with higher antioxidant properties were found to improve the antioxidant capacity of the TMR silage, thereby mitigating the degradation of vitamin A.

Low fertilizer quality and remediation ability are considered the major factors hampering the land application of anaerobic digestate. Therefore, the role of Myrothecium verrucaria treatment in enhancing the fertilizer quality and remediation ability of digestate for land application was explored. Higher content of humic acid (7.5 g/L) with a higher degree of humification index and oxygen-containing functional groups was observed in the digestate receiving Myrothecium verrucaria treatment. Likewise, humic acid formed from Myrothecium verrucaria treatment had a higher capacity of heavy metal binding. Moreover, the viable and culturable count of S. faecalis, S. typhi, C. perfringens, and E. coli pathogens in the digestate decreased to approximately 12.50%, 41.70%, 18.87%, and 50.00% and 25.97%, 64.44%, 37.51%, and 75.27%, respectively, after treatment with Myrothecium verrucaria. This study provided a novel strategy to enhance the fertilizer quality, remediation ability, and biological safety of anaerobic digestate for land application.