Advances in HPLC Stationary phases: Zirconia HPLC columns

Silica-based HPLC columns have remained workhorses in HPLC laboratories. Over the years with advances in particle technologies size reduction has been possible from 5μm to sub- 2μm which led to development of UHPLC. Further introduction of Core Shell Technologies and monolithic columns extended the range of applications and contributed to larger sample throughputs.

Monodisperse, Spherical DIagram copy

Monodisperse, Spherical and Nonporous Zirconia Particles

Properties of Zirconia supports

The main limitation of silica columns is availability of pH range between 2 and 8 and temperature to ambient. On the other hand Zirconia supports exhibits operational stability over the entire pH range and working temperatures up to 200°C.

Lewis acid

Zirconia or ZrO2 is an oxide in which the Zr atom acts as a Lewis acid i.e. – it accepts electron pair due to prescence of empty orbital. This feature provides enhanced selectivity over conventional silica columns.

Adsorption capacity

Adsorption capacity is larger in comparison to silica packings

Chemical resistance

High resistance to acidic and basic solutions. Retention of basic and acidic analytes can be finely controlled by changing the pH of medium over the entire available range.

Types of phases

Four surface chemistries based on surface activities are available which greatly extend selectivity and retention of for improved separations

Polybutadiene coated Zirconia

The phase is similar to reverse phase using C18 silica but has following advantages:

  • Available pH range 1 – 13
  •  Temperature stability up to 200°C
  • Less hydrophobic than C18 and is suitable for separations involving amines and bases

Zirconia-C18

  • Porous carbon clad zirconia particles covalently modified with C18 groups.
  • Selectivity is rather different from conventional C18 columns
  • Can be used over the entire pH range (1 – 14) alongwith high temperature capabilities (up to 200°C)

Zirconia – polystyrene

  • Porous Zirconia particles are modified with cross-linked polystyrene.
  • Tolerate strongly aqueous solutions up (up to 100% pure water)
  • Operational pH range 1 – 13
  • Useful for separation of hydrophobic compounds and amines
  • No requirement for ion pairing requirement
  • Reduced tailing of basic compounds in comparison to silica-based packings
  • Provides mixed mode Reverse phase and Ion exchange separations

Zirconia-Carbon

  • Porous carbon clad Zirconia particles provide enhanced retention of polar compounds
  • Highly hydrophilic surface
  • Useful for separation of isomers and diastereomers
  • Good retention of fused – ring aromatic compounds

Column care

Contaminated columns can deteriorate the peak shape, effect retention times, lead to high backpressure and increase baseline noise. Use of concentrations above 25 mM phosphate, acetate, carbonate/bicarbonate buffers and salts should be avoided. Anions such as carboxylic acids, fluoride and phosphate strongly absorb on Zirconia columns. The recommended steps for column regeneration are

  1. Flush with a 20:80 mixture of acetonitrile and pH 10 ammonium hydroxide for 50 column volumes followed by 10 column volumes of water
  2. Repeat with 20:80 mixture of acetonitrile and 0.1 M nitric acid for same column volumes
  3. Flush with pure methanol, acetonitrile, isopropanol or tetrahydrofuran for 20 column volumes. Zirconia HPLC columns will require the same solvents with at least 20% THF

Storage

  • Zirconia HPLC columns should not be stored in phosphate buffer. The recommended storage procedure is
  • Overnight – 10 volumes of organic modifier/water
  • Long-term storage – 10 volumes of pH 10 ammonium hydroxide followed by 10 volumes of 50: 50 organic modifier/water

We have seen that zirconia-based reverse phases offer a great choice of selectivity and retention. Additional benefits of column chemistries, pH and temperature extremes contribute to versatility of applications. A subsequent article on monolith stationary phase packing will introduce you to further advances in stationary phases.

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About Dr. Deepak Bhanot

Dr Deepak Bhanot is a seasoned professional having nearly 30 years expertise beginning from sales and product support of analytical instruments. After completing his graduation and post graduation from Delhi University and IIT Delhi he went on to Loughborough University of Technology, UK for doctorate research in analytical chemistry. His mission is to develop training programs on analytical techniques and share his experiences with broad spectrum of users ranging from professionals engaged in analytical development and research as well as young enthusiasts fresh from academics who wish to embark upon a career in analytical industry.

Comments

  1. Meinster Kodjo Eduafo says:

    Dear Dr. Deepak

    I think this is a great breakthrough for HPLC analysis. please can send me a quotation for each of them. I will be very grateful. Thank you very much for this insight.

    Best regards

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