Method for measuring extent of microplastic bioaccumulation in bivalves in extreme deep-sea environments

A method for measuring an extent of microplastic bioaccumulation in bivalves in extreme deep-sea environments, comprising the following steps:

• S1, collecting and estimating a total number of bivalves in an area:
• S11, searching for a living bivalve bed in early and middle development stages of methane seeps in deep-sea methane seeps, measuring a bed area, and finding an equivalent diameter and a geometric center of the bed area;
• S12, sampling the bivalve bed several times, with sample positions located at two ends and a midpoint of an equivalent circle radius;
• after sampling and retrieval, counting a number of living bivalves, washing the bivalves, and storing the bivalves frozen after draining water;
• S13, based on a number of bivalves sampled several times, roughly estimating a total number of living bivalves in the entire bivalve bed;
• S2, bivalve morphology measurement:
• thawing sampled bivalve samples, performing morphological data measurements and weighing in batches, and dividing the bivalves according to increasing shell length into four age groups of juvenile, young, middle-aged, and elderly;
• S3, multivariate factor analysis:
• based on obtained morphological data in step S2, associating and analyzing a relationship between bivalves of different ages on seafloor and their external length and wet weight using a multifactor statistical analysis method, and identifying top five individual bivalves representing characteristics of each of the four age groups, resulting in a total of twenty bivalves;
• S4, microplastic enrichment and extraction:
• separating and extracting microplastics from each tissue of representative bivalve samples of the four age groups selected by the multivariate factor analysis in step S3, ultimately obtaining a purified microplastic filter membrane for convenient subsequent observation and detection;
• S5, instrument identification:
• sequentially identifying microplastics with sizes ranging from 0.001-5 mm in the purified microplastic filter membrane obtained in step S4, and comprehensively analyzing an occurrence state of microplastics within bivalve bodies and physical changes of the microplastics after entering deep sea;
• S6, carbon-14 dating method:
• using carbon-14 dating to determine chronological ages of corresponding individual bivalves, thereby converting a lifespan duration of the individual bivalves from the four age groups into time periods of microplastic absorption by seafloor bivalves;
• S7, dynamic microplastic accumulation model for bivalves:
• using an adsorption kinetic model to construct microplastic accumulation curves within bivalve bodies;
• calculating accumulation coefficients to represent a degree of microplastic accumulation in sample bodies relative to an environmental concentration;
• based on a microplastic concentration in the top five individual bivalves representing the group characteristics selected from the four age groups and their corresponding carbon-14 estimated ages, jointly establishing a time-series model;
• extrapolating temporal microplastic adsorption patterns of the individual bivalves obtained from all measurement data to the entire bivalve bed through sampling patterns, and approximately deducing a rate of microplastic accumulation and total microplastic accumulation in an entire seafloor extreme environment ecosystem.