Liquid Chromatography with Mass Spectrometry (LC-MS)

The addition of a mass detector to chromatography systems makes a very powerful tool for the analysis of cyanotoxins. Mass spectra can provide an indication of the elemental composition and structure of an analyte along with determining the quantity of analytes for which reference materials are available with high sensitivity (see Box 14.2). A range of differing systems is available, and very careful consideration is required to determine which fulfils analytical requirements and is within the budget available. Different ion sources are available with positive electrospray ionisation (ESI) most commonly used in the analysis of cyanotoxins. The type of mass analyser (Caixach et al., 2017) also varies and can have a significant impact on cost and the data obtained; hence, it is essential that background evaluation is carried out to ensure the system suits the needs defined during planning (see section 14.1.1). In general, LC-MS will provide data relating to chromatographic retention times, the parent ion masses and fragmentation patterns for each compound as they are eluted. More complex LC-MS/MS systems combine a series of more than one mass detector (e.g., a triple quadrupole mass detector). As analyte ions pass through mass analysers, the former allow the selection of an analyte based on parent ion mass, while the latter allow the selective detection of fragment ions. This makes LC-MS/MS a highly specific analytical technique.

Robust protocols are required for LC-MS/MS as the signal from the MS can be either enhanced or suppressed by matrix interference (salts, organics, etc.). Furthermore, the response (i.e., signal strength relative to analyte amount) can drift over a relatively short time, necessitating a regular and frequent calibration. A routine maintenance protocol is advisable with the interval for cone cleaning determined for different sample matrixes; for example, in studies for the analysis of mussel tissue, cone cleaning was required after 40 samples (Waack, 2017). This interval was determined by spiking an extracted sample and then carrying out repeated, identical sample injections and determining after how many samples the reliability of the detection and quantification diminished.

While LC-MS provides very powerful sensitivity and detection capabilities, the more advanced systems (LC-MS/MS) require a capital investment of around $500 000 USD and an annual running cost of $20-40 000. Furthermore, a high level of staff training is required to use, interpret and maintain these systems, but once established, they provide unrivalled analytical capabilities. It is advisable, where possible, to see one or several systems in operation and have an opportunity to analyse specific samples from the area to be monitored prior to committing to this significant capital investment.

Selecting an Analytical System

The lack of suitable analytical equipment is typically a barrier to monitoring cyanotoxins, and a strong case is often made for capital investment. The influence of current scientific publications frequently draws attention to the significant capabilities of very advanced instrumentation. However, while these systems provide impressive capability, a robust evaluation of the analytical requirement, running costs and infrastructure should be made to inform purchasing decisions. Checklist 14.2 provides some key points to discuss both in-house and with those who have recently invested in cyanotoxin analysis before making decisions. In particular, it is advantageous to develop a regional network, sharing expertise and resources, for example, through a regional centre of competency. This may lead to a decision to use simpler techniques such as ELISA while validating the results periodically by having a small set of samples analysed with advanced techniques elsewhere. Where training is required, it is often more efficient to invite an expert to provide an in-house workshop as this ensures analyses are operational and staff develops confidence using the in-house system. (Note: small grants are often available for this, e.g., through international exchanges and workshop funding.) A further benefit of this approach is continued support from experienced international collaborators ensuring ongoing development of monitoring programmes. Support provided by the system’s vendor generally is charged for. This should be considered in the budget for investment and running costs.

CHECKLIST 14.2 EVALUATING INSTRUMENTAL ANALYTICAL REQUIREMENTS AND SUITABILITY

• What information is needed? Which class of toxins will be the main focus?

Is the main target monitoring compliance of cyanotoxin guideline levels?

  • • Check the cost of consumables, for example, vials, columns, SPE cartridges and solvents in relation to the number of samples expected over time. In many countries, solvents and even high-purity water can be prohibitively expensive (the benefits of ultra-performance liquid chromatography (UPLC) are low flow rates and short runtimes requiring little solvent).
  • • What are the costs of waste solvent disposal required by environmental legislation?
  • • Is the infrastructure appropriate? This includes a stable power supply as fluctuating or intermittent power can rapidly destroy equipment. If not, what are the costs of installing effective power surge protection such as uninterruptible power supply (UPS) systems?
  • • Can room temperature be kept within the range needed by the instruments and analyses (results can be affected by high or fluctuating temperature, so air conditioning is often required)?
  • • Do all purchases include installation and initial training, ensuring that there are available engineers in the area?
  • • What is the cost of a service contract, and is it essential?
  • • For planning to purchase a LC/MS (it requires a nitrogen generator and cannot run efficiently on regular laboratory gas cylinders), ensure that the contract includes either annual service - that is, the cost of service engineers visits - or, if you have technical expertise to carry out the service, the purchase of a service kit.
  • • Talk to users of different instrument manufacturers regarding their experience of service and support especially in your location.
  • • Consider the benefits of partnering with others rather than buying own equipment.
 
Source
< Prev   CONTENTS   Source   Next >