---
title: How do I scale-up my reversed-phase flash chromatography method?
description: This post show a simple way to scale-up reversed-phase flash purification methods.
---

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# [How do I scale-up my reversed-phase flash chromatography method?](https://www.biotage.com/blog/how-do-i-scale-up-my-reversed-phase-flash-chromatography-method)

 Written by [Bob Bickler](https://www.biotage.com/blog/author/bob-bickler) | Jan 23, 2023 2:22:13 PM

Scaling up reversed-phase flash chromatography methods is often necessary as reaction scale increases. This is especially true when other non-chromatographic purification techniques do not work or meet purity and/or yield needs.

While preparative reversed-phase chromatography is an option, this technique suffers from low loading capacity and slow throughput. This post details how to simply scale-up reversed-phase flash purification methods using automated flash chromatography systems and prepacked columns.

For success scaling reversed-phase flash chromatography methods, several operating parameters must be held constant…

- Sample concentration
- Sample load as a percentage of stationary phase mass
- Column media and brand
- Elution solvents
- Elution gradient in terms of column volumes
- Solvent flow velocity ([linear velocity](https://www.biotage.com/blog/how-do-particle-size-and-flow-rate-affect-normal-phase-flash-column-chromatography), not flow rate)
- Detection wavelengths

These parameters are determined during small-scale method development, typically using columns 6 to 60 grams in size. Once a method is optimized on the small column, scale-up is linear based on the ratio of column media masses. For example, if 140 mg of a mixture is successfully purified on a 12-gram C18 column then a 60-gram column will successfully purify 5x that amount, or 700 mg, Figure 1.

Figure 1. Linear scale-up purification of a 7-component mixture. Top - 12 g Biotage Sfar C18 column with a 140 mg load. Bottom - 60 g Biotage Sfar C18 column with a 700 mg load (5x more than the 12 g column).

For this example, the seven previously mentioned operating parameters were held constant, Table below.

| **Parameter ** | **Biotage® Sfär 12 gram** | **Biotage Sfär 60 gram** |
| --- | --- | --- |
| Sample concentration | 350 mg/mL   | 350 mg/mL   |
| Sample load % | 1.17%   | 1.17%   |
| Column brand/media | Biotage Sfär C18   | Biotage Sfär C18   |
| Elution solvents     | MeOH/water | MeOH/water |
| Elution gradient     | 30% MeOH (1 CV)   30-100% MeOH (10 CV) 100% MeOH (2 CV)   | 30% MeOH (1 CV)   30-100% MeOH (10 CV) 100% MeOH (2 CV)   |
| Linear velocity (cm/min) | 9.2   | 9.5 |
| Detection     | DAD UV 190-400 nm   | DAD UV 190-400 nm |

 

By maintaining these seven parameters, purification scale-up is possible even to kilogram scale.

For more information on scale-up please download our whitepaper Optimizing Chromatographic Purification for Rapid Results in Chemical Manufacturing.

[Learn More About Scale-Up](https://www.biotage.com/documents/optimizing-chromatographic-purification-rapid-results-chemical-manufacturing-white-paper)

 

[View full post](https://www.biotage.com/blog/how-do-i-scale-up-my-reversed-phase-flash-chromatography-method)

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