Sampling odours at the whole colony scale

Whole colony volatiles can be collected from colonies using the hive equipment itself as a partial enclosure to concentrate colony volatiles. The colony must be well sealed to capture colony odours before volatiles escape to the outside atmosphere. Either replace leaky hive components or plug the gaps with wax. Collect a sample from air outside the colony as a control since this air replaces the colony headspace.

  1. Add 200 μl of the elution solvent (dichloromethane or hexane) to the filter solvent reservoir (just above the adsorbent packing material). 
  2. Gently push the solvent through the packing material with a clean air or nitrogen flow.
  3. Repeat steps 1 and 2 two times.
    This procedure rinses residual contaminants from the adsorbent filter to prepare the adsorbent filter trap for volatile collection.
  4. Place the filter into a sleeve jacket made of short interlocking sections of rigid Teflon tubing (0.635 cm OD, 0.794 cm OD, 0.952 cm OD) (Fig. 2) to protect the filter from the bees.
    The top of the filter needs to attach tightly to the jacket tubing; otherwise, air will flow around, instead of through, the filter.
  5. Construct a sampling tube out of 0.64 cm OD Teflon tubing that reaches from outside the colony to the centre of the colony (~ 30 cm for a Langstroth deep).
    Cover the end with metal screen to prevent bees from entering the tube.
  6. Carefully insert the sampling tube into the colony either through the entrance or a small hole in the equipment to the centre of the colony.
  7. Attach the adsorbent filter in-line between the sampling tube and a flowmeter-regulated vacuum line leading to the vacuum pump.
  8. Collect colony volatiles by pulling colony headspace through the filter at 600 ml/min to 3l / min with flowmeter-regulated vacuum (exchange at least one volume of colony airspace every 20 minutes). 
    Most volatile collections require 3 to 12 hours to collect sufficient material for GC analysis.
  9. End the volatile collection by removing the filter trap from the vacuum line.
  10. Carefully remove the filter from its protective plastic jacket.
    Secure the filter in a holder.
  11. Add 5 μl of an internal standard solution (80 ng nonyl acetate/µl) directly to the top of the adsorbent packing material with a syringe.
    Avoid touching the packing material with the syringe.
  12. Place a GC vial or vial with a glass insert directly underneath the tip of the filter. 
  13. Add 200 μl solvent (dichloromethane or hexane) to the solvent reservoir above the packing material to extract the trapped volatiles from the filter packing material. 
  14. Gently push the solvent through the packing material at a steady drip with clean air or nitrogen flow.
  15. Cap and store the sample vials in a -80°C freezer until GC analysis (see section 2.2.3.).
  16. Volatile emissions can be calculated by comparing compound peak areas to the known amount of internal standard added to the sample:


Note that quantification may be difficult if large amounts of the whole colony volatiles escaped the colony headspace before collection due to a slow sampling rate (colony volumes/h) or poor seals between hive equipment.

Fig. 2. (a) Assembled and (b) unassembled views of a SuperQ adsorbent filter enclosed in a protective Teflon tube jacket made of interlocking sections of tube.