3.2.1. Setup and volatile sampling

Laboratory collection of headspace odours can be carried out using a static or dynamic sampling method. A detailed description of the merits and disadvantages of these two sampling techniques, adsorbents and their associated desorption techniques have been dealt with in the previous section (see section 2.2.2. on In-situ collection of volatiles). Briefly:

  • Dynamic headspace sampling requires an air supply, an air purification system, flowmeters, quickfit glass containers to hold odour sources with air entry and exit ports, and a mesh screen metallic canister with a tight fitting lid to hold honey bees, copper/Teflon tubing connectors, adsorbents (Super Q, Tenax, and SPME fibres), humidifier and a vacuum supply.
  • Static headspace sampling on the other hand simply requires glass containers with lids fitted with air ports to contain odour sources, SPME adsorbent fibres (suited for collecting both polar and non-polar chemicals) or gas tight glass syringes for sampling head space. Sample the headspace directly with a gas tight syringe by pulling 50 or 100 ml of odour and analyse on a GC or GC-EAD or GC-MS.

An example of a laboratory volatile collection setup for honey bees has been described by Torto et al. (2005). A similar setup is shown in Fig. 4, illustrated using 6 components. In this setup, medical air from a pressurised air tank (not shown in figure) is passed through a copper tubing (component 1) and then through activated charcoal (component 2) to purify it and into to a humidifier containing double distilled water (component – 4). The humidified air is pushed through a y- or t-split (for treatment and control) or multiple ports (manifold) (component 5) to which odour sources enclosed in glass jars are connected to in parallel. The vacuum supply pulls air from the glass jar at a specified flow rate set on the flow meter (component 6) through the adsorbent filters. 

Odour collections can be made from adult worker honey bees only; an entire honey bee comb bearing adult workers, bee brood, pollen and honey; honey comb containing bee larvae; just to name a few depending on the hive odour source of interest and the research question being addressed. However, the experimenter should bear in mind that bee larvae usually become stressed in the absence of worker bees (see sections 2.2.2. and 2.2.3. on in situ volatile collection and analysis).  

Once ready to collect odours in the laboratory and in possession of all the bits and pieces of equipment, following the steps below will ensure a successful process.

  1. To collect odours from worker bees only, gently brush-off workers from a comb into a screen mesh metallic canister containing sugar source (a ball of cotton wool soaked in 50% sucrose solution or sucrose solution in a vial fitted with a dental cotton wick) and close it with its lid once the required population is obtained.
  2. A setup consisting of a canister and sugar source without bees should also be prepared to serve as a control.
  3. Return to the laboratory with the canisters and place them in an appropriate quickfit glass container (e.g. 2 or 5 l glass jar with a tight fitting lid).
  4. To sample representative odours of the hive environment, gently push a honey bee comb or cut out a section of a comb with adult bees, brood, pollen and honey out of its frame.
  5. Place in a clean quickfit glass jar with aeration ports and carry the confined comb and bees back to the laboratory for volatile collection.
  6. Connect one aeration port attached to the glass jar to an air supply and the other to the a vacuum supply.
  7. Pass clean air through the system for 15-20 min to purge out the alarm pheromone produced by the bees and any contaminants before collecting volatiles for a specified period.

 

Fig. 4. (A) Ex-situ volatile collection setup, (B) Adsorbent filter fitted on lid and (C) glass jar with metallic canister containing worker honey bees. (1) copper tubes, (2) air filtering system (e.g. .activated charcoal) , (3) air pressure regulator , (4) humidifier, (5) multiple air supply ports, (6) air flow meters, (7) glass jar containing odour source and 8-air exit monitor (water bubbler which give evidence of air flowing through the setup).

figure4