Introduction

Oryx systems were originally developed for protein crystallization, but are increasingly being used for small molecule and inorganic crystallization. With some small updates to the software and experimental setup, the systems can dispense small molecule and inorganic samples, including experiments with volatile solvents and water sensitive samples.

Robot Hardware

The Oryx system has several features that make it suitable for small molecule crystallization:

Chemically resistant dispensing tips
FEP (fluorinated ethylene propylene) Microtips are used, offering excellent chemical compatibility, similar to PTFE. This allows practically any solvent to be handled without degradation or contamination.

System fluid
The standard system fluid is degassed water. If the sample is not sensitive to water, this can be used without modification. If the sample is water-sensitive, an alternative system fluid can be used. Silicone oil is typically recommended due to its inertness and compatibility with most organic systems.

Plate compatibility
A variety of crystallization plates can be used including non-SBS sized plates with non-standard pitches. Polystyrene, COC or polypropylene plates can be used depending on solvent concentration and compatibility with the plate material. Careful selection is required when working with aggressive solvents to avoid polymer degradation.

Automatic oil dispensing
The system automatically dispenses oil to cover drops. Microbatch-under-oil experiments provide protection from evaporation. This is particularly important for volatile solvents for slower crystal growth over time.

Phase Diagram Experiment Design

The Oryx system enables systematic exploration of crystallization conditions through controlled variation of concentration.

Experiments are typically designed using three components:

• Sample to be crystallized (Green)
• Diluent (Blue)
• Antisolvent (Red)

The ingredients are dispensed simultaneously (or layered) in controlled ratios into each drop.

By varying the proportions, the system samples a range of concentrations across the crystallization phase diagram. The experiment is designed to sample along trajectories that are approximately parallel to the expected solubility curve. This systematic approach makes it much easier to identify conditions that lead to nucleation and controlled crystal growth.

Seeding

Seeding can be incorporated into small molecule crystallization phase diagram experiments to improve reproducibility and crystal quality.

The addition of seed crystals allows experiments to access the metastable region of the phase diagram. In this region:

• Nucleation is suppressed
• Existing crystals can grow in a controlled manner

Crystals formed in the metastable zone typically grow more slowly and with improved morphology compared to those formed under highly supersaturated conditions.

Seeding reduces stochastic nucleation events, enabling more predictable and reproducible outcomes across multiple wells.

Automatic Oiling

Evaporation control is critical in small molecule crystallization, particularly when volatile organic solvents are used. The Oryx system automatically dispenses a layer of oil over each drop immediately after mixing. Commonly used oils include paraffin and silicone oil.

The oil layer:

• Protects from evaporation of solvent
• Provides a barrier to water vapour for hygroscopic solvents
• Maintains stable concentration conditions
• Allows controlled evaporation

Case Study 1. Paracetamol

Paracetamol crystallization can be performed using standard phase diagram exploration. The system enables identification of conditions leading to different crystal morphology through controlled variation of solvent and antisolvent composition.

Case Study 2. Aspirin − Seeded Phase Diagram

Without seeding (above)

• Nucleation may be delayed or inconsistent
• As a result of evaporation of acetone, crystals may form rapidly under high supersaturation (large feather shaped crystal at surface of drop)
• Resulting crystals can be small or poorly formed

With seeding

• Nucleation occurs in a controlled manner
• Crystals grow more slowly at lower levels of super saturation
• Improved size and quality are typically observed

Controlled Crystal Growth Over Time Using Seeding

Time-resolved observation of aspirin crystallization demonstrates:

• Crystal growth at lower saturation when seeds are added
• Seeds crystals grow slowly over time
• Without seeding: rapid nucleation occurs as the drop becomes supersaturated. Fast and irregular crystal growth at the surface of the drop

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