| A lot of nutrient-rich byproducts will be produced during the aquatic product processing,which will cause environmental pollution and waste of resources without effective treatment. Inaddition, ordinary fertilizers contain large amounts of harmful heavy metals, they will enter intosoils with the fertilization process. Cadmium is one of the important pollutants with lastingbio-toxicity in soil, which may migrate and enter the food chains through soil-plant system, anddamage human health. Therefore, this study intends to aquatic byproducts as the carrier, by addingBacillus licheniformis with high capacity of cadmium enrichment to obtain organic liquid fertilizer,which can promote the growth of plants and manage cadmium contaminated soil in dealing withaquatic byproducts simultaneously. It has the practical significance.As aquatic products increased, more and more byproducts are produced, including head,bones, skin and viscera, which are accounted for40-60%of its total weight. In order to improve theutilization of aquatic product processing wastes and the soil pollution of cadmium, in this paper,Katsuwonus pelamis byproducts and B. licheniformis A were selected as experimental materials.The enrichment mechanism of B. licheniformis A was systematically studied. Fish protein organicliquid fertilizer with capability of cadmium enrichment was prepared. The main efforts andconclusions results gained can be expressed as follows:Firstly, Katsuwonus pelamis byproducts were selected as materials in this study. The basicnutrient constituents of byproducts were determined using general methods. The results showedcontent of protein of byproducts was about17.85%±0.15%, and that of crude fat, moisture, ashwere8.24%±0.11%,68.08%±0.76%and5.78%±0.24%, respectively. In the processing ofwaste degradation, with the hydrolysis degree as index, the single factor experiment was adopted todetermine the use of the animal proteolytic enzymes and alkaline protease, and then theBox-Behnken central composite design and response surface method (RSM) were adopted to explore the optimum conditions of the bienzymatic hydrolysis of Katsuwonus pelamis byproducts.It showed that the ratio of the animal proteolytic enzymes and alkaline protease was2:1. Theoptimal bienzymatic hydrolysis conditions were as follows: natural pH, hydrolysis time6h,hydrolysis temperature50.77℃, liquid-to-solid ratio2.24:1, enzyme dosage1.48%. Under suchconditions, the predicted degree of hydrolysis was42.95%, and the validity value was42.43%,which was very closed to the forecast value by RSM. Hydrolysis of Katsuwonus pelamisbyproducts by two enzymes could obtain a higher degree of hydrolysis.Secondly, a cadmium-resisting bacterium, B. licheniformis was acquired after a period ofdomestication. The effects of initial Cd2+concentration, bioaccumulation time, ventilation andinoculation amount on bioaccumulation ability of B. licheniformis were investigated. It showedthat: bioaccumulation can reach a balance when the adsorption time and inoculation amount are24h and6%, respectively. The domesticated bacteria can resist and accumulate Cd2+in the initialmass concentration50-500mg/L.The analysis of FTIR indicated that, with the increase of Cd2+concentration, the number ofFTIR peaks increased. A new added strong absorption peak1734cm-1showed the changes ofcarbonyl and amide (-CO-,-CO-NH-). The typical founctional groups such as-OH,-CO-,-CO-NH-on the cell wall mainly interact with Cd2+.Circular Dichroism Spectra indicated that with the increase of Cd2+concentration, the spectralintensity of positive peaks and negative peaks slightly decreased. Peaks shape and value haveremarkable difference separately, especially a large negative peaks in the210-240nm. Thesesuggested that heavy metal Cd2+has changed the bacterial protein space conformation.Scanning electron microscopic observation and energy spectra showed that under thecondition of Cd2+existing in medium, the average size of bacterial cells varied from1.5-3.0μm×0.75-1.0μm to1.5-5.0μm×0.5-0.75μm (1000mg/L Cd2+). They have a rough surface andcomplexes on the surface. There are more impurities among cells. Much cadmium was detected onthe surface of cells.Transmission electron microscope observations showed that in the presence of cadmium, itssurface is rugged and the body is loose. A large number of particles which form the granular layercould be seen on the cell walls. The density of granular layer increased with increasing Cd2+concentration. Combined with atomic force microscope, particle size is about50nm, presumablycontaining Cd.SDS-PAGE results showed that the protein samples had about20bands. There is nosignificant difference among experiment groups, but some bands as compared with the control group had significant difference. Two-dimensional gel electrophoresis profiles showed: proteinspots of the experimental group (1000mg/L Cd2+) were more than those in control group (withoutCd2+). Protein isoelectric point were distributed in intermediate acidic region. Taking the controlgroup as a reference gel, the software analysis showed that average245protein spots of the controlgroup were detected, and327protein spots in the experimental group. Average matching proteinspots in the experimental group were145, no matching protein spots182, matching rate59%.Selected18differential protein spots were identified by mass spectrometry, the results showed:these differential protein spots had many categories, including energy metabolism (PTS systemglucose-specific transporter subunit IIA), stress response (peroxiredoxin, superoxide dismutase),material transport (trigger factor), transcription (elongation factor Tu) and so on.Thirdly, fish protein organic liquid fertilizer enriched cadmium was made fromconcentrated fish protein degradation liquid, adding spray drying cadmium-tolerant bacteriapowder. Then its various index and cadmium accumulation were studied. The results indicatedthat moisture content of the protein organic liquid fertilizer is40%, the total nutrient content134.2g/L, organic matter content as high as558.3g/L, water insoluble matter29.7g/L, pH5.92.And the content of heavy metals were within the safety range, meeting the enterprise standards offish protein organic liquid fertilizer. At the same time, the fertilizer has high enrichment ability ofheavy metal cadmium. |
