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Gac (Momordica cochinchinensis) oil

Huynh Cang Maia, Frédéric Debasteb

a Department of Chemical Engineering, Nong Lam University, Ho Chi Minh City, Viet Nam. Email: [email protected] Transfers, Interfaces and Processes- Chemical Engineering Unit, Ecole polytechnique de Bruxelles, Université libre de Bruxelles, F.D. Roosevelt street 50, B-1050 Brussels, Belgium. Email: [email protected]

Abstract  

Gac (Momordica cochinchinensis) fruit, originating from South-Eastern Asia, is considered as a su-perfruit thanks to the unequaled content in lycopene and other carotenoids of its arils. Direct uses of the fruit can be considered in cooking or traditional medicine, yet, most interesting and large scale applica-tions in food, cosmetic and pharmaceutical require to extract the gac oil with its carotenoids content. Gac oil production is subject to an increasing attention by the scientific and engineering domain but is still in its infancy compared to other oils productions.

It is proposed to summarize the state of the art of gac oil processing by following the valorization chain. First the fruits properties are presented. Then, the steps used to store the fruit and produce the oil (drying, freezing, oil extraction) are presented. For each step, the different known options are com-pared in terms of process conditions and quality of the resulting oil. The usual properties of the gac oil are then reported with an emphasis on the carotenoid content and anti-oxidant activities. Further pro-cessing of the oils (concentration, carotenoids crystallization) are then addressed. The main existing and foreseen application of gac oils are finally discussed.

1 Gac fruit and its properties

1.1 Origin and name

Gac (Momordica cochinchinensis (Lour) Spreng.) is botanically classified into the family of Cucurbitaceae, the genus of Momordica, and the species of Cochinchi-nensis. This perennial vine was given the name of Muricia cochinchinensis by Lou-reiro, and then Flora cochinchinensis by a Portuguese priest in 1790. In 1826, Sprengel concluded that this plant belonged to the genus of Linné Momordica and changed the name to Momordica cochinchinensis Spreng.(Bailey, 1937). Gac is known for its carotenoid content, which was identified for the first time in 1941 by Guichard and Bui (Vuong et al., 2006). Gac is found throughout the Southeast Asian region from South China to North-eastern Australia. It is not only a traditional fruit in Vietnam but also a native fruit of China, Japan, India, Thailand, Laos, Cambodia, the Philippines, Malaysia and

Bangladesh. Its common names in different countries are presented in Tab. 1 (Vuong and King, 2003). A large, bright-red fruit, gac fruit is known as “sweet gourd” and esteemed as “the fruit from heaven” because of its ability to promote longevity, vitality, and health(Vuong, 2000; Kuhnlein, 2004).

Table 1. Common names of gac in different countries

Language Name Language Name

Latin Momordica cochinchinensis Spreng.Muricia cochinchinensis Lour.Muricia mixta Roxb.

English Spiny bitter gourdSweet gourdCochinchin gourd

Chinese Mu Bie Guo Lao Mak kaoMalay Teruah Thai Fak kaoJapanese Kushika, Mokubetsushi Hindi Hakur, Kakrol

1.2 Culture and production

In Vietnam, gac is cultivated in different regions, from the hills and mountains to the delta and coastal areas. In the Mekong Delta regions of Vietnam, gac grows on dioecious vines and is usually harvested from climbing hedges or wild plants. The vines are generally found on hedges of houses in the province or gardens(Vuong, 2000). Traditionally, in Vietnam gac includes two varieties: gac Nep and gac Te. Gac Nep fruit has bigger size, thicker and darker red aril than gac Te has. There-fore, gac Nep is cultivated more in Vietnam for its size, color and nutritive compo-sition. Nowadays, in Vietnam, gac Nep is being cultivated on an industrial scale to extract oil from the aril for its colorant and healthy benefits. The plant can be grown either from seeds or from root tubers. Humidity, heat, air circulation and light are required for gac seeds germination, which is very sensi-tive to cold and dry condition. Gac seeds are easily germinated in 7-10 days in comfortable conditions. It is cultivated once but harvested for several years. The vines can live up to 15 – 20 years. According to people in the Mekong delta, a gac vine on a frame of 50 m2 can produce from 100 to 200 fruits per year. Gac leaves are alternated and deeply divided from three to five lobes with serrated edges. Gac flowers are pale yellow and solitary in the axils of the leaves (Vuong, 2000). The plant begins to bloom about two months after tubers have been planted. The flow-ers are pollinated by insects. Several vines must be grown together in the vicinity to ensure at least one mature male flower for mature female flowers nearby. Flowering usually occurs in April and continues to August and/or September. It takes about 18 to 20 days for a mature fruit since the emergence of the female flower bud. A plant produces about 30 to 60 fruits weighing 1-3 kg each in its sea-son. The ripen fruit is harvested from August to February (Shadeque and Baruah, 1984).

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1.3 Fruit structure

The fruits of the gac are round or oblong, mature to a size of about 13 cm in length and 10 cm in diameter, densely aculeate, and green when becoming dark orange or red when ripen. Gac fruits are picked when they are the optimal size, weight and color. However, gac fruit is mainly harvested in developmental stages, while gac fruits are orange/red and the seeds are hardened (Bhumsaidon and Chamchong, 2016).Figure 1 shows the morphology of the gac fruit from the outside to the inside in-cluding exocarp, mesocarp, aril, seed. Weight distribution of gac fruit is presented in Tab. 2 (Ishida et al., 2004):

The exocarp of gac fruit is thorny, firstly green and turns orange or red when ripen. It is hard and covered with small spines 4.5 cm in height (Vuong, 2000). In some fruits, the spines are smooth and dense, while the others are hard and widely spaced.

The mesocarp represent nearly 50% of the weight of a gac fruit, which is spongy, orange and 4 cm of thickness (Vuong, 2000). The mesocarp contain aril covering a black or brown seed inside and yellow connective tissue in the middle (Vuong, 2000).

The aril of gac fruit, accounting for 25% of the fruit weight, is red, soft and sticky, 1 to 3 mm of thickness, and is used for cooking. The aril texture is sup-ple and spongy, similar to raw chicken livers. The mesocarp and the aril of gac fruit have slight taste of sweet as a cucumber (Vuong, Dueker and Murphy, 2002). The aril of the gac fruit has high antioxidant activities thanks to its carotenoids content and valuable fatty acids.

The seeds of gac fruit represent about 25% of the weight of the fruits weight. The seeds and arils are prepared with rice for a lustrous appearance and rich in oil, a slight flavor for rice. The seeds of Gac fruits are brown and look like small meteorites with gagged edges and black lines running through them.

Fig. 1 Morphology of the gac fruit

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Table 2 Weight distribution of Gac fruit (Ishida et al., 2004)

Fruit part Fresh weight (FW) (g)

% total FW (%)

Aril 190.0 24.6Seed 130.0 16.8Skin 55.0 7.1Mesocarp 373.7 48.4Connective tissue 22.6 2.9Whole fruit 772.0 100

1.4 Fruit composition

Chemical composition of the fresh gac fruit arils is presented in Tab. 3. The gac fruit aril has a high water content (around 76.8 % fresh weight (FW)). The oil con-tent of the aril is about 17.3 % in dry weight (DW). Variability of the water and oil content is related to maturity, variety and cultivation conditions of gac fruit (Mai, Truong and Debaste, 2013). Some research have also shown that gac fruit contains a protein that can inhibit the proliferation of cancer cells (Chuyen et al., 2015).

Table 3 Chemical compositions of Gac arils (Vuong, Dueker and Murphy, 2002; Mai, Truong and Debaste, 2013)

Results are expressed as mean valuesS.E.M (standard error of the mean), N=3.

The carotenoids content, especially -carotene and lycopene, in the gac aril was found to be much higher than that in other common carotenoid-rich fruit(Aoki et al., 2002; Vuong et al., 2006). Different reports of carotenoids analysis in gac fruit are presented in Table 4. Gac fruit mesocarp has significantly lower carotenoid

Composition Value

Water content (%FW) 76.8± 3.3Oil content (%DW) 17.32.6

Total carotenoids content (TCC, mg/g DW) 6.1±0.2

Crude protein (%) 8.2 0.2

Crude fiber (%) 8.7 1.4

Carbohydrate (g/100g aril) 10.5

Starch (g/100g) 0.14

Pectin (g/100g) 1.25

Cellulose (g/100g) 1.8

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contents than gac fruit arils (Aoki et al., 2002; Ishida et al., 2004). The lycopene content of gac fruit arils is greater than that of other fruits considered to be rich in lycopene such as tomato, watermelon, and guava. The total concentration of ly-copene in mature gac is about 3053 µg / g FW, compared to 40-50 µg / g FW of tomatoes(Ishida et al., 2004). All trans-lycopene is the major pigment in ripen gac fruit and has been studied based on its potential health benefits, bioavailability, and changes that occur dur-ing fruit ripening and subsequent processing (Kubola, Meeso and Siriamornpun, 2013; Müller-Maatsch et al., 2017).The β-carotene content is approximately 10 times higher than in western common vegetables for their rich β -carotene content, such as carrots (Vuong, 2000). The other main carotenoid present is α-tocopherol. The total tocopherol concentration around 76 μg/g tocopherol (FW) (Vien, 1995; Vuong, 2000; Vuong and King, 2003; Vuong et al., 2006).Significant amounts of carotenoids are also present in the fruit mesocarp (Vuong, 2000; Aoki et al., 2002; Vuong and King, 2003; Vuong et al., 2006) and peel (Kubola and Siriamornpun, 2011; Chuyen et al., 2017a). Interestingly, these tis-sues appear to be richer in carotenoids, particularly lutein, prior to maturity (Kubola and Siriamornpun, 2011).

Table 4 Carotenoids concentration in different reports (µg/g FW) (TCC: total carotenoid content (µg/g FW) (extended from (Vuong et al., 2006) )

Reference Mesocarp Aril

β -carotene Lycopene TCC β –carotene Lycopene TCC(West and Poortvliet, 1993)

188,1 891,5

(Vien, 1995) 458(Vuong, 2000) 355(Vuong, Dueker and Murphy, 2002)

175 802

(Aoki et al., 2002) 7-37 0,2-1,6 6-40 60-140 310-460 481(Ishida et al., 2004) 16,3- 58,3 636,2-836,3 1546,5-3053,6 2926(Vuong et al., 2006) 283 83,3±40,4 408,4±178,6 497,4±153,7

The arils and mesocarps of gac fruit contain a significant concentration of fatty acids (from 17% to 22%), which is essential for efficient absorption and transport of β-carotene and other soluble vitamins (Vuong, Dueker and Murphy, 2002; Vuong and King, 2003; Kuhnlein, 2004). The aril oil composition is detailed in Sect. 4.

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2 Fruit storage and preservation approaches

Fresh ripe gac fruit does only conserve for a few weeks after collection : carotenoid content has been shown to decrease fast after 12 days (Bhumsaidon and Chamchong, 2016). Therefore, useful fruit parts conservation is an efficient ap-proach to avoid losses and allow oil extraction or fruit usage during a larger time period (Mai et al., 2013). The two main technics for the fruit conservation are drying and freezing. Freezing of the full fruit as received little attention. Drying of gac aril is the most studied transformation process of gac. Indeed, dried gac fruit aril, plain or in powder form, have their own commercial potential (Tran et al., 2008). Also, drying ease further processing (like pulp separation from seeds (Mai et al., 2013)) and has an huge impact on the yield of oil extraction (Kha, Phan-Tai and Nguyen, 2014) . Most of the developments focus on the aril of gac fruit drying. However, recently, conservation approaches for further valorization of skin (Chuyen et al., 2017a, 2017b) and mesocarp (Trirattanapikul and Phoungchandang, 2016) have emerged.As drying is a thermal process, potentially in presence of oxygen, the key issue of gac aril drying is the preservation of its carotenoids. As the color of the fruit and its antioxidant activity are mostly controlled by the carotenoids content, these pa-rameters show the same global evolution during drying as the carotenoids content (Mai et al., 2013). However, it should be noted that the total anti-oxidant activity evolution can show more complex behavior as thermal isomerisation between dif-ferent carotenoids can have an impact on the anti-oxidant activity for the same to-tal carotenoid content (Phan-Thi and Waché, 2014).Usually, the gac fruit arils are dried to a wet based humidity of 6% at which the water activity is around 0.1 (Tran et al., 2008). Limiting drying to 18% of humid-ity was shown to be a viable option for shorter term conservation (Mai et al., 2013). Peels are dried to a wet based humidity between 2% and 6% (Chuyen et al., 2017a).Five main technics are encountered: oven air drying, vacuum oven drying, heat pump drying spray drying and freeze drying.

2.1. Oven air drying

Gac fruit arils are directly placed in an oven in which hot air flow is achieved. For the carotenoids preservation, optimal drying is achieved around 40°C or 60°C de-pending on the authors (Kha, Nguyen and Roach, 2011; Mai et al., 2013), as a compromised between drying time and degradation of carotenoids. Still, air drying allows only limited conservation of the carotenoids, around 65% at best both for the arils (Kha, Nguyen and Roach, 2011; Mai et al., 2013). Pretreatment by soak-ing with ascorbic acid or bisulfite offer an improvement up to 10% (Kha, Nguyen and Roach, 2011).

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Similar results have been obtained for gac fruit skin drying, with a results less de-pendent on air temperature. An optimal air temperature at 80°C was identified (Chuyen et al., 2017a) to retain 55% of the carotenoid. Pretreatment with as ascor-bic acid allowed to retain 10% more (Chuyen et al., 2017b).Despites a low efficiency in retaining the carotenoids, air drying is widely used due to its simplicity and limited investment costs (Chuyen et al., 2017a).

2.2 Vacuum drying

Vacuum drying, similar to air drying but in a reduced total pressure, allows a faster drying and a contact with less oxygen, limiting the carotenoids decomposi-tions. Up to 90% of the carotenoids from arils could be retained at an optimal tem-perature of 60°C (Tran et al., 2008; Mai et al., 2013). For the vacuum drying of gac skin, no significant improvement compared to air drying was observed (Chuyen et al., 2017a).

2.3 Heat pump drying

Heat pump drying allows efficient lower temperature operation than air of vacuum technics. This approach was only tested for gac fruit skin drying. At 30°C, the re-sults for carotenoids retention were not significantly different from air drying at 60°C due to a longer time of operation (Chuyen et al., 2017a). The interest of in-vesting in heat pump drying for gac fruit skin seems limited.

2.4 Spray drying

Spray drying of gac fruit aril allows the direct production of marketable powder. Direct spray drying of pulp allows only to retain only 6% of the carotenoids (Tran et al., 2008). Optimizing the spray drying, by using an air temperature of 120°C and adding 10% of maltodextrin as encapsulation agent, allows to conserve close to 50% of the carotenoids. (Kha, Nguyen and Roach, 2010). Further optimization using whey gum as carrier (Kha et al., 2014c) and using surface response method-ology for process optimization lead the conservation of 90% of the carotenoids (Kha et al., 2014b) . The achieved powders can be stored during several months with limited loss in their micronutrients content (Kha et al., 2015).

2.5. Freeze drying

Freeze drying is the costliest way to dry gac aril but it offers unequaled carotenoids conservation thanks to its very low temperature and limited contact with air. Different studies show a conservation of carotenoids ranging from 82%

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(Mai et al., 2013), 88% (Bruno et al., 2018) to 100% (Tran et al., 2008). Yet, as this method has high investment and operational costs compared to the previously presented methods, it is seldom used outside of the laboratory for gac fruit drying.

3 Oil extraction

Although the other parts of the fruit potentially contain oil, gac oil refers to the oil contained extracted from arils. Gac oil receives a growing attention as it is the most direct way of accessing the liposoluble carotenoids contained in the arils. Two main approaches can be used to extract oil: mechanical approaches, mostly pressing (Kha et al., 2013), and physico-chemical approaches, based on oil solu-bility in solvents. The two main type of solvent used are organic solvent (Kubola, Meeso and Siriamornpun, 2013) and supercritical fluids (Tai and Kim, 2014). Yet, despites oil not being soluble in water, enzymatically assisted water extraction is also possible (Mai, Truong and Debaste, 2013; Thi, Nhi and Tuan, 2016).For all the technics, various pretreatment of the arils can be considered to enhance the extraction efficiency such as drying (Kubola, Meeso and Siriamornpun, 2013; Kha, Phan-Tai and Nguyen, 2014), microwave heating coupled with steaming (Kha et al., 2013), ohmic heating (Aamir and Jittanit, 2017) or enzymatic treat-ment (Kha et al., 2013; Mai, Truong and Debaste, 2013; Thi, Nhi and Tuan, 2016). These treatment, through heat of enzymatic activity affect gac aril cell structure, easing oil extraction.The various pretreatment and extraction methods can have a varied impact on the oil content, including on the heat sensitive carotenoids (Kha et al., 2013).

3.1 Mechanical extraction

Pressing is the most common extraction method encountered. While low cost and technologically simple, it usually allows to extract only around 70% of the avail-able oil. For gac oil, this yield can be achieved with air dried arils with 170 kg/m². Combining microwave drying (at 630W for 65 min) and partial re-humidification by steaming, it was possible to reach 93% of yield. This treatment seems to offer a better conservation of carotenoids present in the oil, doubling the amount com-pared to air dried aril oil (Kha et al., 2013).

3.2 Organic solvents extraction

Various organic solvent have been tested for gac oil extraction: chloroform/ methanol mix, petroleum ether (Kubola, Meeso and Siriamornpun, 2013), n-hex-

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ane (Thuat, 2010; Kubola, Meeso and Siriamornpun, 2013; Aamir and Jittanit, 2017), About 95% of the oil contained in air dried gac aril can be extracted with hexane at 50°C but the process can take up to 18h (Thuat, 2010). Treating the dried arils with commercial enzymatic mixtures Viscozyme L (composed of arabanase, cellu-lase, hemicellulase, β-glucanase and xylanase) can reduce the extraction time to 2h. Fresh aril submitted to ohmic heating during the extraction exhibit yields close to 100% (Aamir and Jittanit, 2017).On top of hazardous question raised by organic solvent, the use of gac oil for food and pharmaceutical applications tend to reduce the interest of organic solvent ex-traction that have to be totally removed from the final product before consump-tion.

3.3 Supercritical fluid extraction

Supercritical fluid extraction combine the efficiency of the organic solvent extrac-tion with the ease of separation of the supercritical fluid from the extracted oil. On top of that, the extraction is usually faster than with organic solvents. The main opposition to the use of supercritical fluid come from the investment cost and the requirement of a specifically skilled workforce of this advanced technology (Mar-tins, de Melo and Silva, 2015). Supercritical carbon dioxide is the most common supercritical fluid and is the only one tested for gac oil extraction as well as for a specific extraction of carotenoids (Kha, Phan-Tai and Nguyen, 2014). As for other extraction methods, the pretreatment applied to the gac aril is has a drastic impact on the amount of oil that is effectively extracted. Supercritical car-bon dioxide at 200bar and 50°C flowing at 70 kg.h-1.kggac

-1 allowed to extract 95% of the total oil content of 50°C air dried arils in 3 hours. An additional treatment with 0.1% of pectinase enzymes allowed to achieve the same yield in only 2 hours (Kha, Phan-Tai and Nguyen, 2014). However, the most efficient way to accelerate the extraction is rise the pressure: 95% of the oil of air dried aril can be achieved in 30 minutes at 400 bar (Tai and Kim, 2014).

3.4 Enzymatic aqueous extraction

Using enzyme to free the oil contained in gac aril is a low investment alternative cost to solvent extraction. A high yield of oil extraction requires the synergic effect of multiple enzymes. A mix of pectinase, cellulase, protease and α-amylase allowed to extract 82% in slightly more than 2h (Mai, Truong and Debaste, 2013). However, the operational cost of the enzymes acquisition can hinder the profitability of such a process.

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4 Oil properties

The main properties of the extracted oil depends on the oil extraction process and upstream processing as well as on the initial fruit that is used and its storage con-ditions (Bhumsaidon and Chamchong, 2016).

4.1 Fatty acid composition

Table 5 summarizes the main fatty acid present in gac oil as obtained by different authors with various extraction methods. Variations are observed between the studies. They can be attributed to difference in fruit initial content, extraction tech-nics as well as quantification methods. However, global trends can be highlighted.The majority (56% to 75%) of the fatty acid present in gac oil are unsaturated with mainly oleic acid. Most of the rest of the unsaturated fatty acid are poly-unsatu-rated with a dominance of linoleic acid. In this fraction, significant concentration in ω-3 fatty acid are present, mainly in the form α-linolenic acid. In the saturated fatty acid, palmitic acid is the most present. The oil also contains a significant amount of stearic acid. The mixture of unsaturated, saturated, poly-unsaturated and mono-unsaturated fatty acids in gac oil improves the absorption and bioavailability of nutrients and carotenoids(Müller-Maatsch et al., 2017).

Table 5 Fatty acid composition of gac oil for different methods of extraction

Study (Vuong, Dueker and Murphy, 2002) (Thuat, 2010)

(Kha et al., 2014a)

(Thi, Nhi and Tuan, 2016)

(Mai, Truong and Debaste, 2013)

(Kha et al., 2014a) (Kha et al., 2014a) (Bruno et al., 2018)

Extraction method Organic solvent Organic solvent Organic solvent Enzyme and water Enzyme and water Press Microwave and press Supercritical CO2

Lauric (C12:0) 0.02 0.02

Myristic (C14:0) 0.87 0.21 1.09 0.37 0.22 0.63 0.41 0.8

Pentadecanoic (C15:0) 0.1

Palmitic (C16:0) 22.04 20.27 34.73 24.18 17.31 34.89 24.99 30.1

Palmitoleic (C16:1) 0.26 0.23 0.19 0.16 0.18 0.18 0.4

Margaric (C17:0) 0.23 0.15 0.14

Stearic (C18:0) 7.06 5.35 8.45 3.52 7.45 7.78 6.85 5.1

Oleic (C18:1) 35.21 49.57 45.04 48.99 59.5 40.58 48.25 44.5

Linoleic (C18:2) 31.43 23.19 10.14 21.09 13.98 15.6 18.28 19.6

α-linolenic (C18:3) 2.14 0.94 0.37 0.86 0.52 0.34 0.83

Arachidic (C20:0) 0,39 0.21 0.32

Eicosa-11-enoic (C20:1) 0,15 0.23 0.17

Arachinodic(C20:4) 0.1

Docosanoic (C22:0) 0.19 0.03

Erucic (C22:1) 0.07 0.1

Docosahexanoic (C22:6) 0.02

Tetracosanoic (C24:0) 0.14 0.04

4.2. Physicochemical properties

Table 6 summarizes typical physicochemical properties of gac oil including non-saponification matter, refraction, melting point, viscosity and density value (Mai, Truong and Debaste, 2016). The iodine value of gac aril oil corresponds to a high degree of unsaturated oil (76g I2/100g oil). The high saponification value (715 mg KOH/g) indicates that the triglycerides of gac oil are composed of short fatty acids.Table 7 presents acidity value and peroxide values depending on the treatment. The acidity value of gac oil is between 0.69 and 3.6 mg KOH/g depending on the extraction method, which is lower than that of some common oils such as soybean oil (about 6 mg KOH/g). The lowest peroxide value measured (around 0.89 meqO2/kg oil) characterizes the purity and stability of this oil at ambient tempera-ture.

Table 6 Physicochemical properties of the Gac oil (Mai, Truong and Debaste, 2016)

Index Value

Iodine value (gI2/100g oil) 76.58±1.9Saponification value (mg KOH/g) 715.16Non-saponification matter (%) 0.5Refraction value (n25D) 1.47Melting point (°C) 12Viscosity (Pa.s) 0.0466 ±0.0004Density (g/ml) 0.955 ±0.012Results are expressed as mean values S.E.M (standard error of the mean), N=3

Table 7 Acidity value and peroxide values depending on the treatment

Authors TreatmentAcidity value (mg KOH/g)

Peroxide (meqO2/kg oil)

(Thuat, 2010) Hexane extraction 3.58 8.7

(Kha et al., 2014a) Microwave + press 0.69 1.8

(Kha et al., 2014a) Press 1.8 7.7

(Kha et al., 2014a) Solvent extraction 2.19 33.54(Mai, Truong and Debaste, 2013) Enzymatic extraction 2.55 0.89

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4.3 Carotenoids content and other minor bioactive compounds

As for other properties, fruits and processing variation have a significant impact on the carotenoid content. As carotenoids are thermosensitive molecule, the pro-cessing conditions have an exacerbate impact on the carotenoids content. On top of that, multiple studies have shown qualitatively that, for any given treat-ment, rising the extraction yields also leads to an increase of the retrieved carotenoids concentration (Kha et al., 2013; Mai, Truong and Debaste, 2013; Tai and Kim, 2014; Thi, Nhi and Tuan, 2016). This can probably be attributed to the fact that the carotenoids are preferably stored in gac fruit regions that are more dif-ficult to reach.

Table 8 summarizes the total carotenoid content, lycopene and β- carotene con-centrations obtained by various author with different processing. The total carotenoids content in the gac oil can go up to 10 mg/g, in which lycopene and β-carotene are representing a majority, up to 5 mg/g for each.

Table 8 Mass concentration (in mg per g of oil) of total carotenoids, lycopene and β-carotene using various processes

Authors TreatmentTotal carote-noids (mg/g)

Lycopene (mg/g) β-carotene (mg/g)

(Thuat, 2010) Hexane 3.2(Thi, Nhi and Tuan, 2016) Hexane + enzymes 7.7(Aamir and Jittanit, 2017) Ohmic heating + hexane 1.5 5.8(Kubola, Meeso and Siriamornpun, 2013)

Chloroform + methanol ex-traction 0.49 1.18

(Mai, Truong and De-baste, 2014) Enzymatic extraction 7.9 3.4 2.69

(Kha et al., 2014a) Press 2.51 0.57

(Kha et al., 2014a) Microwave + press 4.33 1.46(Kha, Phan-Tai and Nguyen, 2014) Supercritical CO2 5.08 0.83

(Tai and Kim, 2014) Supercritical CO2 10.9

(Bruno et al., 2018) Supercritical CO2 4.63 2.4 1.57

These amounts in carotenoids are larger than what is reported in usual fruit and vegetables. Common source of lycopene compound includes tomato (31 g/g), watermelon (41g/g), guava (54g/g) and pink grapefruit (33.6 g/g) (Mangels et al., 1993; Aoki et al., 2002).Carotenoids predominantly occur in their all-trans configuration, which is ther-mos-dynamically the most stable isomer. All-trans-lycopene may be converted to

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cis configuration during processing and they possess different biological proper-ties (Lee and Chen, 2002). Several reports have demonstrated that the cis-isomers of lycopene are absorbed into the body more easily and play a more important part in biological function than all-trans-lycopene (Böhm and Bitsch, 1999; Failla, Chitchumroonchokchai and Ishida, 2008).cis-isomers of lycopene correspond to 2.7 to 13.2 % of the total while cis-isomer of -carotene range between 6.1 and 25.3% of the total .The-carotene was also found at a lower concentration (1% of the total carotenoids) in gac oil (Ishida et al., 2004; Bruno et al., 2018). Moderate heat treatment (exposure of oil at 80°C for 4h) lead to isomerization of lycopene contained in the oil from a fraction of all-trans-isomer to different cis-isomer, mainly 13-cis (22% of the total) and 9-cis (16%) isomers (Phan-Thi and Waché, 2014).Carotenoids extracted from natural source are usually in the free form or as fatty acid esters. Zeaxanthin and -cryptoxanthin have hydroxyl groups in the 6-membered ring, and these hydroxyl groups can bind with fatty acids to form carotenoid esters. There-fore, zeaxanthin and -cryptoxanthin were found in saponified oil samples. The degree of esterification of carotenoids in some fruit increases during ripening (Subagio and Morita, 1997).The analy-sis of the saponified samples detected also a trace of zeaxanthin and -cryptoxanthin (Aoki et al., 2002).

4.3 Antioxidant activity

The antioxidant capacity of gac oil strongly depends on the con-tent and bioavailability of carotenoids, especially, β-carotene and lycopene (Mai et al., 2013) like for other oils (Thaipong et al., 2006). Other phenolic compounds, vitamin E and unsaturated fatty acids including oleic and linoleic acid, contribute also on antioxidant ca-pacity of gac oil. Lycopene exhibits a high physical quenching rate of singlet oxy-gen (Di Mascio, Kaiser and Sies, 1989; Perretti et al., 2013), which is directly related to its antioxidant activity. The antioxi-dant activity of lycopene in multi-lamellar liposomes is superior to other lipophilic natural antioxidants (e.g. α-tocopherol, α -carotene, β-cryptoxanthin, zeaxanthin, β -carotene and lutein) (Stahl and Sies, 2007).

While the existence of a significant antioxidant activity of gac oil is generally well accepted, their quantification leads to potentially contradictory results in the few existing studies.

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Most studies do not directly measure the antioxidant activity of the oil but rather of dry powder (Kha, Nguyen and Roach, 2010, 2011; Mai et al., 2013). Then, some hydrophilic compounds also play a role in the antioxidant activity (Mai et al., 2013). The comparison of the antioxidant properties in various conditions is also hard-ened by the discrepancies between the methods used to evaluate the antioxidant activity. Moreover, the different process have complex impacts on these activities. Indeed, while treatment induce can induce a loss in carotenoids, moderate heat leads to isomerization that can raise the antioxidant activity (Phan-Thi and Waché, 2014).

With the ABTS antioxidant assay (Thaipong et al., 2006), it was shown that the global antioxidant activity is lower with the drying at higher temperatures, in the range of 0.3 mmol Trolox equivalent per gram of powder (TEAC) at 40°C to 0.2 TEAC at 80°C (Kha, Nguyen and Roach, 2011). This seems contradictory, or at least underlying more complex phenomena, with a study showing a doubling of the TEAC with a treatment of oil at 80°C during 4 h (Phan-Thi and Waché, 2014).With the DPPH test (Thaipong et al., 2006), one study highlight similar result as with ABTS, with a reduction of the activity from 0.25 TEAC to 0.2 TEAC (Kha, Nguyen and Roach, 2011) while another suggest that the activity is best conserved at 60°C where 0.18 TEAC (80% of the activity observed for the fresh sample) would be conserved. The same study shows results going accordingly with FRAP while DMPD tests gave uncorrelated results (Mai et al., 2013).

5 Oil concentration and carotenoids extractions

Further transformation of the oil can be considered to enhance its carotenoids con-tent or to valorize pure carotenoids. Few works have dealt with that question. Crossflow filtration of gac oil with a cut-off size of 5nm was shown to allow a concentration of total carotenoids in the retentate of a factor 8.6 while the acid in-dex was divided by a factor 40 (Mai, Truong and Debaste, 2014).Crystallization of the carotenoids by mixing the oil with propylene glycol fol-lowed by saponification with KOH, allows to achieved crystals containing 94% of carotenoids (Mai, Truong and Debaste, 2016). Other processes could be considered for oil concentration. An example of poten-tially interesting development would be the use of supercritical carbon dioxide, which was proved to efficiently extract the oil to fractionate the oil, like it was done for tomato oil (Perretti et al., 2013).

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6 Existing and foreseen gac oil product

Gac oil is a premier source of carotenoids, especially beta-carotene and lycopene. Because of high content of carotenoids and fatty acid and its high bioactivity, gac fruit can be consumed as a natural treatment for vitamin A deficiency in children in developing countries (Vuong, Dueker and Murphy, 2002; Vuong and King, 2003). Gac oil can be used in many food applications, for example, cooking oil, salad oil, seasoning and food coloring, or in cosmetic applications including soap and skin oils.

6.1 Food applications

In Vietnam, only the aril of ripen gac fruit is traditionally used as natural colorant and additive for cooking. For example, it is added into sticky rice to produce a brilliant orange rice dish known as “xoi gac”. This meal is prepared by mixing araculae of gac fruit with cooked rice to give a red color, a lustrous appearance and a distinct flavor. This food preparation is served as one of special meals at New Year celebration, wedding and other important celebrations. Consumption of this traditional food could produce a substantial increase in -carotene intake (Vuong, 2000). In addition, women and children in Vietnam readily accepted consumption of gac oil because it can help reducing lard intake (Vuong, Dueker and Murphy, 2002; Vuong and King, 2003). In Thailand, Gac aril is cooked and eaten with chili paste or cooked in a curry (Kubola, Meeso and Siriamornpun, 2013). The incorporation of 1.0% gac aril powder can, therefore, be used to reduce the amount of nitrite added to Vienna sausage from 125 ppm to 75 ppm, resulting in more red and darker sausage with higher lycopene content (Wimontham and Rojanakorn, 2016). Gac oil is used as an additional nutrition and natural food additives. β-carotene is well known for its pro-vitamin A activity. Lycopene is added to foods to increase the nutritional value of products, in particular for dairy products, energy drinks and fruit juices. β -carotene can also be added to livestock feeds to improve the quality of milk or eggs. The orange to red colors of lycopene and β-carotene are widely used in foods and beverages as natural colors, which are usually consid-ered to be safer than other artificial colors. These natural colors are used to en-hance, change or contribute to the color of food products such as fruit juices, sweets, butters, cheeses and sauces. In addition, they can act as antioxidants to ex-tend product shelf life.

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6.2 Non-food applications

Gac oil is also used as a traditional medicine. For example, gac fruit has been ap-plied to treat conditions of the eyes, burns, skin problems and wounds. It was re-ported that when applied to wounds, skin infections and burns, gac oil stimulates the growth of new skin and the healing of wounds. The fruit is also frequently used as a traditional remedy for arthritis and cardiovascular diseases and degener-ation of the macula (Burke, Smidt and Vuong, 2005).Because of its high concentration of beta-carotene, gac fruit is a valuable aid in preventing or treating vitamin A deficiency. Therefore, the gac aril is used to make a tonic for children and lactating or pregnant women, and to treat xeroph-thalmia and night blindness (Guichard and Bui, 1941). In many developing countries, vitamin A deficiency is epidemic because it can lead to poor night vision, blindness, higher rates of maternal mortality, poor em-bryonic growth, and reduce ability to fight infections and lactation. Supplementa-tion with gac fruit extract can alleviate chronic vitamin A deficiency, and help re-ducing these health problems (West and Poortvliet, 1993; Vuong, 2000).The lycopene and β-carotene in gac fruit enhance skin health by mitigating oxida-tive damage in tissue. The various antioxidants in gac fruit boost heart health by specifically combating atherosclerosis. Additionally, both lycopene and beta-carotene show protective activity against the risk of heart attack.Gac oil could be used as pharmaceutical ingredients: pure lycopene and β-carotene are commonly used in pharmaceuticals, such as multiple vitamin tablets for a nu-tritional supplement or for vitamin A deficiency patients.Several new gac products are currently being developed on the world market. All products obtained from gac have antioxidant characteristics, determined by the bioactive compounds it contains, such as lycopene, -carotene and vitamin C.In Vietnam, Vnpofood is the largest manufacturer with a capacity of 3000 tons of gac fruit per year. Their products include gac oil capsules (Vinaga). These gac oil capsules contain pure gac oil. Another brand of gac oil capsules is Garotene, pro-duced by the University of Hanoi Pharmacy. These capsules are enriched with vi-tamin E. There is also a product called G3, produced by Pharmanex, a company in the United States. It is a combination of lycium chinensis fruit phytonutrients, Siberian pineapple and gac fruit.

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