HPLC analysis of microplastics involves sample preparation (digestion/separation), extraction with organic solvents, and HPLC analysis using a C18 column with gradient elution. Key steps include calibration with standards, UV-Vis detection, and data analysis comparing retention times and peak areas to identify and quantify plastic compounds.

To conduct HPLC analysis for microplastics in fish tissues and sediment samples:

1- Prepare samples by digesting organic matter, separating microplastics by density, and extracting them using solvents.

2- Analyze microplastics using HPLC with a C18 column, UV-Vis or MS detection, and calibration standards for quantification.

Solution of KOH or H2O2 can dissolve the tissue leaving out the microplastics. Solvents such as TFA, chloroform and phenol can help to dissolve common variants of microplastics. These extractives can be analyzed using C18 column. However, depending on the composition of microplastics using only one type of solvent will not work and furthermore strong organic solvents will damage HPLC column.

A better strategy would be deconstructing the plastics and dissolving the monomers in benign solvents. For example, PET (a common microplastic) can be hydrolyzed in HCL or NaOH media and its monomer (TPA) can be dissolved completely in DMSO. It can be filtered and injected into the c18 column for analysis. A recommended HPLC condition for this analysis is gradient flow of 0.1% or 1% formic acid and acetonitrile at 0.6 m/L and elevated column temp. A calibration plot using external std concentrations is needed for quantification. Details can be found in the following source: http://dx.doi.org/10.1021/acssuschemeng.3c08286

Lastly, other techniques such GPC-MS, Py-GC-MS, and FTIR can help augment results from HPLC analysis.

Column Selection: Use a reverse-phase HPLC column, typically a C18 column, known for its versatility in separating non-polar to moderately polar compounds. The column should be capable of withstanding the mobile phase conditions, typically involving organic solvents like methanol or acetonitrile, which help in the separation of additives or degradation products from microplastics. The column length and particle size (e.g., 5 µm) should be chosen based on the resolution required, with a length of 150-250 mm being common for adequate separation. Ensure the column’s compatibility with UV or MS detection depending on the analysis needs.

microplastics cannot be analysed by HPLC-UV. Check the current literature.

Isolate the MPs, run either IR or Raman microscopy