SEC-MALS

• SEC-MALS: Overview

Size exclusion chromatography with multi-angle light scattering is used for the determination of absolute molecular weight (MW). A concentration detector, such as refractive index (RI) or ultraviolet (UV), is required for all SEC and SEC-MALS applications. Classical SEC uses RI or UV detection and polymeric standards to establish the retention times of polymers with known MW to create a calibration curve. This is traditionally used to estimate the MW of samples, but can produce errors when the standards do not accurately represent the shape, size, or interactions of your samples with the column's stationary phase. The combination of SEC with MALs uses the relationship between light scattering and molar mass (Rayleigh ratio), which allows the determination of the absolute molecular weight without the use of standards from the equation below.

Lab Overview and Guidelines

In this lab, each student will work to determine the absolute MW of polystyrene or other polymer of choice (with approval), including MW distributions (Mn, Mw, Mz, and D). Traditional SEC-UV calibration will be created to estimate MW and compare to MALS absolute MW determinations. Batch determination of dn/dc will be performed to give the most accurate MW calculations. An interim report (PowerPoint) will be generated after completing the dn/dc lab to evaluate performance before continuing with the SEC-MALS lab. After the SEC-MALS lab, a compiled final report (PowerPoint) will be submitted.

• SEC-MALS Lab Guidelines

Prelab Assignment and Standard Operating Procedures

The prelab assignment must be completed prior to beginning Lab 1 and will be reviewed for any knowledge gaps. The SOPs should be reviewed prior to beginning either lab and should be followed accordingly. The laboratory instructor/operator will help as needed, but a basic understanding and direction should be understood.

• Prelab Assignment

• Batch dn/dc SOP

• SEC-UV-MALS-RI SOP

• ASTRA Data Analysis Summary

Lab 1: Determine dn/dc

The dn/dc value refers to the change in refractive index per the change in concentration. This value is required for the Rayleigh ratio to be completed and to determine an accurate MW. Outside of dn/dc, all other values in the Rayleigh ratio can be determined allowing the absolute MW to be calculated. Thus, this value is of extreme importance and should be calculated precisely.   

• Batch dn/dc SOP

Lab 2: SEC-MALS

SEC-MALS analysis will be performed on a polymer of choice. Using the intensity of light scattering and the dn/dc value determined in Lab 1, the absolute MW will be determined. This value will be compared to the estimated value from traditional SEC-UV calculations using polystyrene standards and their retention times to estimate MW.  

• SEC-UV-MALS-RI SOP

Video Tutorials

• MALS Data Processing

• Conventional Column Calibration (UV Calibration)

• Conventional Column Calibration Analysis (UV Analysis)

• Import Multiple Peaks Into Calibration