Rewriting the Book on Supercritical Fluid Chromatography

References 1. Gros Q, Duval J, West C, Lesellier E. On-line supercritical fluid extraction-supercritical fluid chromatography (SFE-SFC) at a glance: A coupling story. TrAC Trends in Analytical Chemistry. 2021;144:116433. doi:10.1016/j.trac.2021.116433 2. Communications to the editor. The Journal of Organic Chemistry. 1962;27(2):700-706. doi:10.1021/jo01049a069 3. Zosel K; Studiengesellschaft Kohle mbH. Process for the decaffeination of coffee. United States patent 4260639A. 1970. Rewriting the Book on Supercritical Fluid Chromatography Back to basics: SFC and supercritical CO2 Fluids become supercritical when they are highly compressed and take on the properties of both a liquid and a gas. Supercritical carbon dioxide (CO2 ) can be used as the mobile phase in supercritical fluid chromatography (SFC). Until recently, this technique had been overlooked in favor of high-performance liquid chromatography (HPLC) and gas chromatography (GC). However, new developments in the field have demonstrated that SFC can be a highly cost-effective, accurate and sensitive solution capable of outperforming HPLC and GC. In light of these new technologies, it’s time to reshape the field of chromatography with SFC. A supercritical state occurs when a gas or liquid is subjected to temperatures and pressures that exceed its critical point. In this state, it will display properties of both liquids and gases, the advantages of which can be applied to SFC: Using supercritical fluids for chromatography is not a novel idea. In fact, supercritical fluids have been used as a mobile phase for separation for over 60 years, emerging originally under the name “high-pressure gas chromatography”. Rapid carrying and eluting for an efficient mobile phase Improved solvent capabilities High flow rate and variable solvation strength Increased density comparable to a liquid Diffusivity comparable to a gas Dynamic viscosity comparable to a gas Supercritical extraction and chromatography has been used in industrial applications for decades, beginning with the decaffeination of coffee in the 1970s.3 HPGC was initially performed using supercritical monochlorodifluoromethane and dichlorofluoromethane, before carbon dioxide (CO2 ) was discovered as a potential mobile phase.1, 2 H H F F O O F Cl C C C Cl Cl Why CO2 ? CO2 has several advantages over other supercritical fluids, making it the standard mobile phase for high-performance SFC. CO2 Phase Diagram CRITICAL TEMPERATURE = 31°C CRITICAL POINT SOLID SUPERCRITICAL FLUID CRITICAL PRESSURE = 7.3 MPA LIQUID GAS SUBCRITICAL FLUID TEMPERATURE (°C) PRESSURE (MPA) Critical temperature and pressure can be achieved without environmental extremes Reverts to a gas at room temperature and atmospheric pressure, resulting in high recovery rates and rapid purifications Miscible with most co-solvents, allowing dissolution of a wide range of both polar and non-polar analytes Non-toxic and non-corrosive Inexpensive and abundant at high purity Chemically inert CO2 Time to rethink supercritical fluid chromatography Over its lifetime, SFC has often been overshadowed by other chromatography techniques. As a result, many first-time users of SFC may have misconceptions about the modern face of this technique, but it’s time to set the record straight. Misconception Fact SFC is only useful for chiral molecules CO2 -based SFC can only be used for non-polar compounds SFC requires specialized expertise and equipment Detection isn’t sensitive enough for many applications SFC is expensive and timeconsuming SFC can be used to separate both chiral and achiral molecules and is often used to separate pharmaceutically active compounds such as vitamins, steroids and barbiturates. Adding polar solvents allows SFC to separate polar substances (e.g. small peptides, polar pesticides, etc.). Furthermore, the supercritical properties of CO2 make it well suited to comprehensively analyze compounds over a wide polarity range not possible with HPLC or GC alone. The same principles of chromatography apply to both HPLC and SFC, meaning that expertise is fully transferable. Additionally, the columns (e.g., silica and octadecyl) and detectors (e.g., photodiode array (PDA) and mass spectrometry) used in HPLC can be directly transferred to SFC. Coupling SFC with a mass spectrometer (MS) can enable greater sensitivity in some circumstances than LC/MS. This is due to an alternative ionization mechanism caused by the interaction of CO2 with the solvent. SFC systems have a higher benefit-to-cost ratio compared to HPLC, as the low viscosity and high diffusivity of supercritical CO2 allow for higher flow rates for faster analysis and separation, with no loss of sensitivity. Chiral Achiral Suitable for a wide range of applications SFC is a well-established technique in the pharmaceutical industry. However, it is also suitable for a wide variety of industries and applications: • Purification of drugs • Analysis of drug formulations • Chiral separations • Pharmacokinetic studies • Forensic applications • Lipidomics • Bioactivity analysis in traditional medicine • Quality control and contaminant analysis • Production of sustainable natural products • Contaminant analysis • Quality control • Analysis of complex compounds, e.g., lipids, pesticides PHARMACEUTICS BIOANALYSIS NATURAL PRODUCTS HIGHLY VERSATILE CHROMATOGRAPHY: NEXERA UC AND NEXERA UC PREP FOOD SCIENCE End-to-end solutions with Shimadzu Shimadzu has created a comprehensive “Unified Chromatography” analytical scale system, in Nexera™ UC; but what is Unified Chromatography? UC characterizes the use of a very wide gradient, from a CO2 -rich supercritical/ sub-critical mobile phase (SFC) to a completely liquid mobile phase like in LC.1 This is a versatile technique that allows separation of hydrophobic and hydrophilic compounds in a single analysis. To meet current needs for a reliable preparative scale instrument, Shimadzu has worked closely with the Enabling Technologies Consortium (ETC), a group of pharmaceutical and biotechnology companies, and released the Nexera™ UC Prep. This system offers the high performance of the Nexera UC with reliable high-performance semi-prep purification. Both systems also enable the on-line extraction (SFE) and analysis (SFC) of a wide range of target samples for high-sensitivity detection and/ or purification. Analytical scale UHPLC/SFC switching system allows screening by UHPLC and SFC in same batch 94.2% reduction in organic solvent consumption for improved sustainability Fully automated on-line extraction and separation Fractionation/purification at both analytical and preparative scales with addition of a fraction collector High resolution even at high flow rates Meeting every need: Shim-pack UC columns Due to the gas-like properties of the SFC mobile phase, diffusion of the sample band is higher than in liquid chromatography. Although standard HPLC columns can be used for SFC, higher speed and performance can be achieved with dedicated columns. Separation of the same three analytes in different columns shows that retention behavior differs significantly according to the column chosen. Shimadzu's Shim-Pack UC series offers a wide range of column sizes (analytical and preparative scale), and packing materials, to ensure a solution for almost every analyte. AQbD methods can significantly reduce the number of data points needed for optimization – by up to 52% Improving efficiency: LabSolutions™ MD software To reduce time-consuming and intimidating method optimization processes, Shimadzu has developed the LabSolutions™ MD software. By configuring parameters, including mobile phases and columns, analysis schedules are automatically generated, and optimal analytical conditions are determined. This allows for simpler, more efficient and beginner-friendly experimental design. Make analysis schedule based on experimental design Analysis with automated switching of co-solvents, columns and SFC parameters Building of Design Space with chromatogram simulation Transform your chromatography with SFC SELECT COLUMNS OPTIMIZE YOUR WORKFLOW Nexera UC UFMS System Nexera UC Prep - Stacked Fraction System Advantages of CO2 HPLC Column: Shim-pack HRC-Sil (250 x 4.6 mm I.D.) Mobile phase: N-hexane/2-propanol (99/1) Flow rate: 1.0 mL/min Temperature: 40°C Detection: UV 290 nm SFC Column: Shim-pack UC-Sil (250 x 4.6 mm I.D.) Mobile phase: CO2 /Methanol (95/5) Flow rate: 3.5 mL/min Temperature: 40°C Back-pressure: 10 MPa Detection: UV 290 nm