|
" Development, Chemometric-Assisted Optimization and In-House Validation of a Modified Pressurized Hot Water Extraction (PHWE) Method for Multi-Mycotoxin Analysis "
Gbashi, Sefater
Njobeh, P. B.
| Document Type
|
:
|
Latin Dissertation
|
| Language of Document
|
:
|
English
|
| Record Number
|
:
|
1112329
|
| Doc. No
|
:
|
TLpq2526189784
|
| Main Entry
|
:
|
Gbashi, Sefater
|
|
|
:
|
Njobeh, P. B.
|
| Title & Author
|
:
|
Development, Chemometric-Assisted Optimization and In-House Validation of a Modified Pressurized Hot Water Extraction (PHWE) Method for Multi-Mycotoxin Analysis\ Gbashi, SefaterNjobeh, P. B.
|
| College
|
:
|
University of Johannesburg (South Africa)
|
| Date
|
:
|
2019
|
| student score
|
:
|
2019
|
| Degree
|
:
|
D.Tech.
|
| Page No
|
:
|
268
|
| Abstract
|
:
|
Improvement in analytical methods, including extraction procedures has been a recurrent topic in mycotoxin research due to the fact that mycotoxins are ubiquitous, potent and notoriously difficult to eradicate from the food and feed supply chain. Effective management of these toxicants rely on stringent regulation and routine surveillance of food commodities via efficient analysis, hence the continuous need for improved methods. This research work describes for the first time the optimization, validation and application of a modified pressurized hot water extraction (PHWE) method for the simultaneous extraction of multiple mycotoxins and subsequent quantification on ultra-high performance liquid chromatography coupled to a tandem mass spectrometer (UHPLCMS/MS). The research was performed in three phases which are presented as chapters (i.e., Chapters Three to Five), in this thesis. The first part of the research aimed to establish the thermal degradation patterns of mycotoxins and determine their relative stability under different temperature and time conditions. This was performed in order to determine the feasibility of the application of PHWE for the extraction of the mycotoxins since this extraction technique involves the use of heat. Using a set of statistically pre-determined experimental conditions (103 to 217 °C, and 6.72 to 63.28 min for temperature and time, respectively), simultaneous optimization of the thermal degradation of 15 different mycotoxins in pure form and when spiked into maize was achieved. The mycotoxins studied included aflatoxins (AFs), fumonisins (FBs), zearalenone and its analogues α-zearalenol and βzearalenol (ZEAs), ochratoxins (OTs), T-2 toxin (T-2), alternariol monomethyl ether (AME) and sterigmatocystin (STEG). The global optimum degradation condition (i.e., minimum temperature and time required for complete degradation of all mycotoxins) was computed as 217 °C/63.28 min and 211 °C/54.71 min for pure mycotoxin standards and spiked maize matrix, respectively. Based on the results obtained, it was inferred that the studied mycotoxins demonstrated sufficient thermal stability to undergo PHWE which typically employs temperatures ranging from 50 to 200 °C and extraction times ranging from 5 to 20 min. The second part of this research work focused on the development, optimization and validation of a modified PHWE method for the simultaneous extraction of multi-mycotoxins from maize. This part of the research was aimed at providing technical solutions to some eminent challenges of conventional methods of mycotoxin extraction, such as, use of large volumes of harmful organic solvents, high cost, long and tedious procedures as well as, as the inherent challenges of simultaneously extracting multiple mycotoxins in a single step. The adoption of advanced chemometric optimization models coupled with a co-solvent-modification of the PHWE system proved vital to achieve satisfactory recoveries across the analytes. The recovery values ranged from 71 to 124%, with the exception of ochratoxin A (OTA) and α-ZEL which had recoveries of 58 and 14%, respectively. These values were comparable to those of at least three other methods which are solvent-based, validated and well referenced in literature. The linearity (0.986 to 0.999), intra-day precision (0 to 27%) and inter-day precision (3 to 34%) values of the modified PHWE method were also good relative to European Commission (EC), Codex Alimentarius (CODEX) and the Association of Official Analytical Chemists (AOAC) guidelines for analytical method performance. Furthermore, a pilot-scale application of the method on 25 maize flour samples purportedly naturally contaminated with mycotoxins was successful. The third phase of this research work describes the broader application of the modified PHWE method for the estimation of mycotoxin levels in different commercially available food commodities. Ninety-one (91) samples of maize, sorghum and millet obtained from 5 agroecological zones in Nigeria were analyzed for mycotoxins using the modified PHWE method. For further authentication, the same samples were analyzed using a validated solvent extraction method which served as a control for cross-validation. Scrutiny of the results using principal component analysis (PCA) and orthogonal projections to latent structures discriminate analysis (OPLS-DA) indicated that there were no significantly different patterns in the data obtained using the two methods. Accordingly, the results revealed that all of the maize samples (n=16), 32% (n=38) of sorghum and 35% (n=37) of millet samples were positive for at least one of the 15 tested mycotoxins. Amongst the toxins, fumonisins (FBs) were the most prevalent in terms of incidence rate and contamination levels. Aflatoxin (AF) contamination was also relatively high in the samples. In maize, mean contamination levels for aflatoxin B1 (AFB1) and sum of aflatoxins (ΣAFs) was 54 and 56 µg/kg, respectively. Eleven out of 16 of the maize samples were contaminated above the EC maximum level of 2 and 4 µg/kg for AFB1 and ΣAFs, respectively. Generally, mycotoxins were more prevalent in maize, followed by sorghum and then millet. Cooccurrence of two or more mycotoxins, particularly AFs+FBs, were observed in some of the cereal samples, which re-affirmed the seriousness of the mycotoxin issue in Nigeria and the need for effective and sustainable intervention approaches to address this issue. Overall, PHWE seems promising as a suitable and greener alternative to traditional methods of mycotoxin extraction. Research in this area is envisaged to allow bioanalytical scientists working in the field of mycotoxicology to conveniently and efficiently extract mycotoxins in various agricultural products without the need for harmful and expensive organic solvents, as well as, promote analytical techniques that are more environmentally friendly and sustainable.
|
| Subject
|
:
|
Accuracy
|
|
|
:
|
Calibration
|
|
|
:
|
Discriminant analysis
|
|
|
:
|
Economic impact
|
|
|
:
|
Food contamination poisoning
|
|
|
:
|
Food science
|
| |