Liquid chromatography-mass spectrometry analysis of compounds from complex matrices

Bowerbank, Samantha (2021) Liquid chromatography-mass spectrometry analysis of compounds from complex matrices. Doctoral thesis, Northumbria University.

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Abstract

The aim of the work was to establish two methods for the analysis of 4 thyroid hormones and 18 plant toxins by liquid chromatography-tandem mass spectrometry (LC-MS/MS) using electrospray ionisation (ESI). The developed and validated methods were then applied to samples, with a range of complex matrices, namely serum, simulated gastro-intestinal fluid, tablet formulation and plants, with sample preparation methods developed and validated for each.
Thyroid disorders, hypothyroidism and hyperthyroidism, are a common medical condition with diagnosis obtained via the analysis of thyroid hormones present within serum samples. Currently within the NHS analysis is performed using electrochemiluminescence to measure thyroid stimulating hormone, thyroxine and triiodothyronine. However, these methods have limited sensitivity and inaccuracies of 30% at the limit of detection. Therefore, diagnosis of hypothyroidism can often be missed with multiple medical visits being required. The developed method allows for the detection and qualification of 4 thyroid hormones, thyroxine, triiodothyronine, reverse triiodothyronine and diiodothyronine, using low resolution and high resolution mass spectrometry. The developed method showed significant improvements over the current NHS method with LOQs of <5.9 pmol/L, r2 of >0.990 and precision of <3.8 %. A direct comparison between the development method and the current clinical method showed that the result obtained were comparable in terms of thyroid hormone levels detected. Allowing for LC-MS/MS to be a suitable, more reliable alternative for the detection and quantification of thyroid hormones.
Plant toxins are a common cause of accidental poisoning with a range of symptoms including burns, nausea, convulsions, paralysis and even death. Many of the plants analysed in this study can be purchased from garden centres and online nurseries with little to no safety information provided. A method was developed for the analysis of 18 plant toxins, atropine, aconitine, capsaicin, cathinone, colchicine, coumarin, digitoxin, digoxin, hellebrin, 5-methoxypsoralen, 8-methpxypsoralen, psoralen, salvinorin A, scopolamine, solanine, strychnine, thujone and veratridine, based upon the plant availability. The developed method showed good linearity, r2 > 0.9952 and precision of <10.7 %. The analysis of the plant materials showed toxins consistent with information obtained from the head gardener at Alnwick Gardens, which holds the largest poison garden in the UK. A risk assessment for ingestion and dermal exposure was compared with the LD50 and it was found that Colchicum autumnale posed the greatest risk via ingestion with Atropa belladonna also posing significant risk via ingestion. The dermal risk was minimal; however, care should be taken in the case of dermal contact due to the presence of phototoxins present in plant such as Heracleum mantegazzianum.

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