The roots used in the study came from KwaZulu-Natal, South Africa, and were authenticated at the University of South Africa under voucher specimen number KGM081118. The researchers ground the dried roots into a fine powder, sieved it through a one-millimetre mesh, and extracted the oil once, using hydrodistillation in a Clevenger-type apparatus with a five-litre round bottom flask for five hours. The yield was 0.89 percent by weight of the starting plant material. The recovered oil was light brown and carried a vanilla-ginger-like scent, an aromatic signature the authors connect to one particular aldehyde that dominated the profile.

That dominant compound was 2-hydroxy-4-methoxybenzaldehyde, which accounted for 20.93 percent of the oil in the one-dimensional analysis and 18.96 percent in the two-dimensional dataset. The compound has previously been reported to display antimicrobial, antioxidant and antibacterial activities in laboratory studies, and earlier work has suggested it is responsible for the characteristic aroma of the plant’s roots. The authors argue, however, that the esters scattered throughout the oil also contribute to the scent, given how widely distributed they proved to be. Other abundant constituents included hexadecanoic acid ethyl ester, m-guaiacol, octanoic acid ethyl ester, and 3-hydroxy-4-methoxybenzaldehyde, alongside hexadecanoic acid methyl ester and octadecanoic acid ethyl ester among the major esters.

Class by class, the oil was dominated by esters. In the one-dimensional analysis, half of the identified compounds were esters when methyl 8-oxooctanoate, which straddles the ester and ketone categories, was excluded, and esters contributed 31.42 percent of the oil by percentage area, with phenolics close behind at 29.73 percent. Ketones and fatty acids each made up 10.53 percent of the identified compounds by number. The two-dimensional method sharpened this picture considerably: seventeen additional esters were found, bringing that class to 53.26 percent of the total oil content, while the number of detected ketones rose from four to nine. Only one fatty acid, hexanoic acid, appeared in the two-dimensional run, and several compounds seen by the conventional method were too small to register there because their peaks were suppressed by larger neighbours.

The technical heart of the study lies in how the two instruments differ. In one-dimensional gas chromatography-mass spectrometry, compounds are separated on a single medium-polar column before mass detection. The comprehensive two-dimensional setup instead passes the eluent through a second column of different selectivity, in this case a non-polar phase, with a modulator and secondary oven coordinating the transfer. Because analytes are separated on two columns with different selectivities, closely eluting compounds that would merge into a single peak on one column can be resolved in the second dimension. The authors note that simply lengthening a single column cannot achieve the same result for compounds of similar volatility, and it would prolong the analysis time.

A concrete example illustrates the gain. Ethyl oleate and (E)-9-octadecenoic acid ethyl ester are isomers sharing the molecular formula C20H38O2. On the medium-polar column of the one-dimensional instrument they were not sufficiently resolved and appeared as a single peak, so only ethyl oleate was identified. Under the two-dimensional method the two isomers separated cleanly: ethyl oleate eluted at 1470 seconds in the first dimension and 1.78 seconds in the second, while its isomer eluted at 1464 seconds and 2.19 seconds respectively. The same technique also distinguished two isomers of tetradecanoic acid ethyl ester, with first-dimension retention times of 1172 and 1596 seconds and second-dimension times of 0.994 and 0.167 seconds.

Every compound assignment in the study is explicitly tentative. The authors classified identifications by confidence, requiring similarity and reverse match scores above 800 together with probability values above 1000 for higher confidence, and they stress that definitive identification would demand confirmation with authentic reference standards. Compounds failing at least two criteria, or scoring below 600 on similarity, were labelled low confidence, while those missing one criterion or lacking a literature retention index were rated moderate. Retention indices were calculated against a C8 to C40 alkane standard and compared with literature values from NIST and NCBI databases. The oil was analysed after one week of storage at four degrees Celsius, diluted in GC-grade hexane at ten milligrams per millilitre, injected split-less in one-microlitre volumes, and run in triplicate with solvent blanks before and between injections to guard against contamination and carryover.

The chemical inventory also carried a cautionary note. Several phthalic acid esters, chemicals used as plasticizers and additives that are considered potential hazards to ecosystems and public health, were tentatively identified in the oil, along with 2,4-dinitrophenyl crotonate, a derivative of a pesticide reported in soils. The authors suggest such compounds may enter plants through human activities, including agricultural pesticide use and industrial emissions. Several polyaromatic hydrocarbons, including phenanthrene, fluoranthene and a methylated phenanthrene, were also detected. The researchers are careful to state that the identities and concentrations of these contaminants require confirmation through targeted analyses before any conclusions about potential exposure or health risk can be drawn.

Because fatty acid esters made up most of the identified esters, the authors connect the profile to the plant’s reported biological activities, noting that fatty acid esters have been reported to possess antimicrobial activity and that the major esters have individually been linked in the literature to anticancer, antibacterial and anti-inflammatory properties. They are explicit, however, that targeted confirmation and bioactivity-guided assays are required before any biological effect can be assigned to individual constituents of the oil. The study also places its findings in geographic context: earlier work on Nigerian material identified twenty-eight compounds dominated by (E)-2-hexen-1-ol, heptacosane and phytol, while an analysis of oil from Cameroon reported menth-1-en-7-al as the main compound, a variation the authors attribute to differing environmental conditions and plant age rather than to any change over time.

For a plant whose roots have been used for centuries across West, Central, East and Southern Africa, and whose extracts have drawn research attention for potential therapeutic relevance, the chemical baseline supplied here is a methodological advance as much as a phytochemical one. By demonstrating that comprehensive two-dimensional gas chromatography with high-resolution mass spectrometry can lift the fraction of the oil that can be profiled from roughly seventy-two percent to ninety-seven and a half percent, the study sets a template for future work on complex essential oils. The authors frame their contribution as an addition to the phytochemistry of Mondia whitei and suggest that the compounds identified, some of which have documented activities in prior laboratory studies, may warrant further investigation of their potential contribution to the medicinal properties reported for the plant, always pending the confirmatory analyses their tentative identifications still require.

Subject of Research: Chemical profiling of the essential oil from Mondia whitei roots using GC-MS and comprehensive two-dimensional GC-HRT-MS.

Article Title: Analysis of essential oil from Mondia whitei (Hook. f.) Skeels roots using GC-HRT-MS and GCxGC-HRT-MS

Article References: Chokwe, R. C., Dube, S., & Nindi, M. M. (2026). Analysis of essential oil from Mondia whitei (Hook. f.) Skeels roots using GC-HRT-MS and GCxGC-HRT-MS. Heliyon, 12(16), Article e45609. https://doi.org/10.1016/j.heliyon.2026.e45609

Image Credits: AI Generated

DOI: 10.1016/j.heliyon.2026.e45609

Keywords: Mondia whitei, essential oil, GCxGC-HRT-MS, gas chromatography, mass spectrometry, 2-hydroxy-4-methoxybenzaldehyde, esters, phytochemistry, tentative identification, phthalates, KwaZulu-Natal, hydrodistillation