Profiling of volatile flavor compounds in nkui (a Cameroonian food) by solid phase extraction and 2D gas chromatography time of flight mass spectrometry (SPME‐GC×GC‐TOF‐MS)

Abstract The objective of this study was to investigate the volatile flavor compounds of nkui, a Cameroonian food, using solid phase microextraction (SPME) and a two‐dimensional gas chromatography time of flight mass spectrometry GC×GC‐TOF‐MS system. Using SPME, volatile compounds were extracted from nkui and analyzed by GC×GC‐TOF‐MS. The data retrieved revealed the presence of flavor volatiles including acids (20%), alcohols (4%), aldehydes (10%), aromatic compounds (4%), esters (7%), furans (4%), ketones (11%), terpenes and terpernoids (27%). Although the terpene compounds were the most predominant, an ester (linalyl acetate) had the highest percentage of 19%, conferring a sweet, green and citrus flavor. Results obtained from this study suggest that the characteristic flavor of nkui was due to the combination of different volatile flavor compounds, which contributed to its aroma. Considering the medicinal importance of these compounds, their presence positions nkui as a vital food source with health benefits and medicinal properties.


| INTRODUC TI ON
Plant parts are frequently used as spices, food additives, and meals.
Nkui is a traditional heavily spiced Cameroonian soup, consumed as food and utilized in traditional medicine for nursing mothers.
Flavor is a major factor that determines consumer selection, perception, acceptance of a particular food product and thus plays a significant role in the food. The flavor of any food is thus largely dependent on the number, quantity, and characteristics of the different volatile compounds it contains (Jelen, Majcher, & Dziadas, 2012).
Such is extended to food substances and subsequent preparations from them. Despite the rich flavor and other volatile components present in nkui, previous studies that investigated nkui for these compounds have not been presented in the literature.

O R I G I N A L R E S E A R C H
Profiling of volatile flavor compounds in nkui (a Cameroonian food) by solid phase extraction and 2D gas chromatography time of flight mass spectrometry (SPME-GC×GC-TOF-MS) extraction (SBSE), pressured hot water (PHW), and liquid-liquid extraction (LLE) are known (Gbashi, Adebo, Piater, Madala, & Njobeh, 2017;Goncalves et al., 2016), challenges around cost, environmental friendliness, ease of extraction, and cross-contamination are their major challenges. The emergence of solid phase microextraction (SPME) has provided an efficient, robust, selective, cost effective, and environmentally friendly extraction technique and their combination with an effective detection technique offer enormous potentials (George et al., 2018;Kusano, Kobayashi, Iizuka, Fukushima, & Saito, 2016). The effectiveness of SPME is also demonstrated in its ability to integrate extraction, concentration, and analyte injection into a single process, ensuring sample throughput (Goncalves et al., 2016).
GC×GC-TOF-MS is sensitive chromatographic technique for both separation and detection of compounds. Its capabilities with includes increased mass spectral identification and deconvolution algorithms, identification abilities, enabled detection of thousands of peaks and the provision of additional data makes it a suitable analytical platform. Its compatibility with SPME for profiling of volatile flavor compounds has been reported (Ding, Wu, Huang, & Zhou, 2016;Goncalves et al., 2016). The aim of this study was thus to investigate the flavor components in nkui using SPME and subsequent analysis on GC×GC-TOF-MS.

| Nkui sampling and composition
Different plant materials that make up nkui were collected, identified by the National Herbarium in Cameroon, and deposited as specimens. The specimens were issued voucher numbers and the different plant parts and quantities used to make up the nkui are presented in Table 1.

| SPME sampling
The extraction of nkui flavor components was done using a 50/30 μm SPME fiber coated with divinylbenzene/carboxen/polydimethylsiloxane (DVB/CAR/PDMS) (Supelco, Inc., Bellefonte, PA). Briefly, 20 g of the sample was placed in a head-space vial heated at 40°C for 20 min. Sampling was then done by exposing the fiber to the headspace of the sample for 20 min, after which the SPME device was transferred to the GC×GC-TOF-MS equipment for analysis. The process was repeated four times, though before each analysis the fibers were thermally cleaned and conditioned by heating them at 270°C in a stream of helium. A SPME fiber (Supleco, South Africa) was used to sample the volatile compound and immediately injected into a GC-MS system for analysis. as the secondary column (Restek, Bellefonte, PA, USA). Helium was used as a carrier gas at a constant flow rate of 1 ml/min and an inlet temperature of 250°C. An initial oven temperature of 40°C was set and held for 0.5 min and then slowly ramped at 10°C/min to 250°C and then held for 0.5 min at 250°C. The modulator and secondary oven were run at an offset temperature of 5°C above the primary oven. The mass spectrometer was set up under the following conditions: no solvent delay because it was a SPME analysis; transfer line temperature at 250°C; Electron ionization at −70 eV; source temperature at 250°C; stored mass range: 45-600 μ; acquisition rate: 10 spectra/s for GC×GC-TOF-MS; detector offset voltage was set at 300 V.

| GC×GC-TOF-MS analysis
Retention time alignment, matched filtration, peak detection, and peak matching were done on ChromaTOF software (LECO, USA). Subsequent identification was done by comparison with mass spectral databases (NIST, Adams, and EO libraries). A semi quantification of each compound was calculated on the basis of peak areas and relative concentration presented in %.
According to Takahashi, Sumitani, Inada, and Mori (2002), aldehydes could be formed through the degradation of polyunsaturated fatty acids (PUFA), either by enzymatic action or autoxidation and lipid peroxidation. Few aldehyde compounds were observed in this study (Table 2), with the most significant in terms of % occurrence being 3,7-dimethyl-2,6-octadienal, with a lemon odor. The presence of 3,7-dimethyl-2,6-octadienal has also been reported in several plants and fruits (Burdock, 2010). Similarly, ketone compounds with different chain lengths are abundant in nature and largely contribute to the flavors in plants and spices. Terpenes and their epoxides could possibly react to form ketones and alcohols (Preedy, 2009).
Seven esters were identified in this study with linalyl acetate and terpinyl propionate being the predominant ones, with 19% and 1.5%, respectively, found (Table 2). Esters are characterized by pleasant fruit odors that contribute to the aromatic, sweet, and honey notes in foods (Burdock, 2010). Linalyl acetate ( Figure 1) is a naturally occurring phytochemical commonly found in spices and plants. It is an ester of linalool having a sweet, green, and citrus smell ( anti-inflammatory, anti-hypertensive, and analgesic effects (Peana et al., 2002).

ACK N OWLED G M ENTS
The authors acknowledge Mr. Alexander Whaley of LECO Africa for kind assistance with the use of their equipment and application laboratory.

E TH I C A L R E V I E W
This study does not involve any human or animal testing.

CO N FLI C T O F I NTE R E S T
The authors declare that they do not have any conflict of interest.