Effect of cold-press and soxhlet extraction on fatty acids, tocopherols and sterol contents of the Moringa seed oils

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Effect of cold-press and soxhlet extraction on fatty acids, tocopherols

and sterol contents of the Moringa seed oils

M.M. Özcan

a

, K. Ghafoor

b,

⁎

, F. Al Juhaimi

b

, I.A.M. Ahmed

b

, E.E. Babiker

b

a

Department of Food Engineering, Faculty of Agriculture, Selcuk University, 42031 Konya, Turkey

b_{Department of Food Science & Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia}

a b s t r a c t

a r t i c l e i n f o

Article history: Received 12 March 2019

Received in revised form 25 April 2019 Accepted 11 May 2019

Available online 6 June 2019 Edited by AR Ndhlala

The current study presents physico-chemical attributes, fatty acid, sterol and tocopherol compositions in oils ob-tained from Moringa seed using either cold-press or Soxhlet extraction systems. Moringa oleifera and M. peregrina seed oils, recovered using cold press (CP) and Soxhlet extraction (SE) techniques, contained substantial amount of oleic acid (75.49 and 74.62% in CP 73.83 and 71.67% in SE) showing signiﬁcant (p b .05) differences. Palmitic acid contents of M. oleifera and M. peregrina oils were 9.58 and 11.74% in case of CP and 12.27 and 12.84% in SE. Theα-tocopherol contents of M. oleifera and M. peregrina oils obtained using these two techniques were 139.61 and 137.89 mg/kg in CP and 142.74 and 151.67 mg/kg in SE systems. Stigmasterol contents of M. oleifera and M. peregrina oils obtained with cold press and Soxhlet extraction system were found as 19.62 and 17.84% to 18.54 and 15.41%, respectively.

Keywords: Moringa oleifera Moringa peregrina Oil Chemical properties Fatty acids Tocopherols Sterols GC–MS HPLC 1. Introduction

Moringa spp., from Moringaceae family, is a widely distributed plant species (Lalas and Tsaknis, 2002; Ashraf and Gilani, 2007). Moringa is considered as a useful plant in tropical and subtropical re-gions and different parts (ﬂower, fruits and roots) of this plant are considered edible (Rahman et al. 2009; Abd El Baky and El-Baroty, 2013). The fatty acid composition (except linoleic) of Moringa seed oil is similar to olive oil, and considered as an acceptable substitute of the former (Lalas and Tsaknis, 2002). Moringa seeds have been used as a culinary oil in the South cost of Red Sea region (Abd El Baky and El-Baroty, 2013). At the same time, the Moringa oil demon-strated a strong antifungal activity against a zoophilic dermatophyte (Chuang et al., 2007). Moringa seeds are rich in oil known as“Ben oil” or “Behen oil” (Palafaox et al., 2012; AL Juhaimi et al., 2016a). The ben oil has been used in salads (Tsaknis et al., 1999). The oil pro-duction usually incorporates the use of three methods which are pressing technique, Soxhlet system and a combined use of pre-pressing followed by extraction using solvents (Özcan et al., 2013).

Moringa seed oil obtained using cold press system can be used as ed-ible oil due to the possibility that it may contain higher proportions of essential fatty acid and other functional compounds in comparison to some other edible oils. Due to the fact that it does not involve use of heat and organic solvents, cold press oil can be regarded as safe and more natural (Goldberg, 2003). The recent trend and increase in consumption of cold pressed vegetative oils can be attributed to better nutritional properties. Cold-press process also has several advantages over solvent-based systems, such as being environment friendly and requiring lesser energy (Rotkiewicz et al., 1999). Despite being the fact that cold pressing can yield better quality edible oil from plant seed, solvent-based extraction systems still have wider applications in food industry and this may be due the lack of comparative studies involving use of two techniques for recovery of Moringa seed oil. In current study, experimental results of Soxhlet (using petroleum ether) and cold-press extraction systems are reported. The aim of this study was to investigate the effects of cold pressing and Soxhlet extraction on physico-chemical properties, fatty acids, tocopherols and sterol contents of oil obtained from M. oleifera and M. peregrina.

⁎ Corresponding author.

E-mail address:kghafoor@ksu.edu.sa(K. Ghafoor).

https://doi.org/10.1016/j.sajb.2019.05.010

Contents lists available atScienceDirect

South African Journal of Botany

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2. Material and methods 2.1. Material

The dried Moringa (Moringa oleifera and Moringa peregrina) seeds were obtained from Saudi Arabia and Sudan. Seeds were cleaned and sieved to remove any debris and unwanted material andﬁnally ground to a powder form using mortar grinder. The powdered seed samples were kept in 4 °C till prior to extraction of oil using Soxhlet system. Whole seed were used for extraction of oil using cold press.

2.2. Method 2.2.1. Cold pressing

The Moringa seed oil recovery using cold pressing was carried out using seed which were cleaned to remove broken or damaged seed and other unwanted materials such as skin, leaves and stem. The cold press system (2–6 l/h capacity) was operated without using heat. The recovered oil was clarified using one week sedimentation process as im-purities can negatively affect oil composition and quality followed by fil-tration. The clarified pure oil was kept in colored bottles which were hermetically capped and sealed and stored under the nitrogen at +4 °C. 2.2.2. Soxhlet extraction

The soxhlet extraction was carried out from powdered seed samples using petroleum ether and the temperature was set at 50 °C. The Soxhlet apparatus was operated for 6 h to recover the oil followed by removal of solvent in rotary evaporator. The residue oil was kept in colored vials at −18 °C till analysis.

2.2.3. Physico-chemical properties

Different physico-chemical properties of oil samples, including acid, peroxide, density, iodine, refractive index, saponiﬁable matter and unsaponiﬁable matter values, were evaluated using StandardAOAC (1990)methods.

2.2.4. Fatty acid composition

The analytical procedure for determining contents of different fatty acid in Moringa seed oil involved their esterification as explained inISO-5509 (1978)method. The analysis was carried out using a gas chromatography (Shimadzu GC-2010) system that consisted of a flame-ionization detector (FID) and capillary column (Tecnocroma TR-CN100, 60 m × 0.25 mm,film thickness: 0.20 μm). Nitrogen was used mobile phase and itsflow rate was set at 1.51 ml/min (total flow rate being 80 ml/min with a split rate of 1/40) whereas the injection block and detector temperature was kept at 260 °C. The programming of col-umn temperature included 120 °C for 5 min which was increased to 240 °C @ 4 °C/min and held for 25 min at this temperature (AOAC, 1990). 2.2.5. Tocopherol content

The analytical procedure for estimation of tocopherols in Moringa oil samples involved use of chromatographic procedure in which a solution oil was prepared by dissolving 250 mg oil sample in 25 ml of n-Heptane. The HPLC system consisted of a Merck-Hitachi low-pressure gradient system which was equipped with an L-6000 pump, a Merck-Hitachi F-1000fluorescence detector and a D-2500 integration system. The detec-tor was set 295 nm excitation wavelength whereas the emission wave-length was 330 nm. The injection volume was 20μl and it was maintained using a Merck 655-A40 auto-sampler. The HPLC column used was a Diol phase column having dimensions of 25 cm × 4.6 mm ID (Merck, Darmstadt, Germany). Theflow rate was set at 1.3 ml/min. n-Heptane and tert-butyl methyl ether in a ratio of 99/1 (v/v) was used as mobile phase. Standard solutions ofα, β, γ and δ-tocopherols, formulated using 0–100 mg/l standard concentration, were used for quantification purposes (Balz et al., 1992).

2.2.6. Sterol analysis

The determination of sterol involved saponification of 250 mg oil sample using a solution of ethanolic potassium hydroxide by boiling under reflux. The unsaponifiable matter was eluted by flushing the col-umn using 20 ml ethyl ether and ethyl acetate (1:1) mixture. The separation of sterol and unsaponi_{fiable fraction was carried out} using thin-layer chromatography (TLC) which was followed by re-extraction from the TLC material. The analysis of sterol fraction was then carried out GC against an internal standard, the botulin and chro-matographic evaluation was carried out on a GC (HP5890) instrument that contained a SE 54 CB fused silica capillary column (Macherey-Nagel, Düren, Germany; 50 m long, 0.32 mm ID, 0.25μm film thickness). The sample volume was set as 1μl and delivered to the system using an auto-sampler. A split ratio of 1:20 was used during analysis. Mass spec-trophotometer temperatures were set as MS transfer line at 260, MS ion source at 220 °C and MS quadruple at 160 °C. The ionization energy was 70 eV (Matthäus and Özcan, 2006).

2.3. Statistical analyses

The statistical analyses were carried out using analysis of variance techniques using JMP version 9.0. The analytical measurements were carried out in triplicates and the results were reported as mean ± stan-dard deviation (MSTAT C) of independent Moringa samples (Püskülcü andİkiz, 1989).

3. Results and discussion

Physical and chemical properties of Moringa oleifera and M. peregrina oils extracted using either cold pressing or Soxhlet extraction systems are presented inTable 1. The cold pressed oil samples' acid values were quanti_{ﬁed as 0.09 (M. oleifera) and 0.07 mg KOH/g (M. peregrina),} acid values of M. oleifera and M. peregrina seed oils obtained with Soxhlet extraction were determined as 0.14 and 0.11 mg KOH/g, respec-tively. In addition, peroxide values of M. oleifera and M. peregrina ex-tracted with cold press and Soxhlet extraction system were determined 0.18 and 0.16 meqO2/kg to 0.35 and 0.27 meqO2/kg,

respec-tively. Also, while unsaponifiable matter amounts of M. oleifera and M. peregrina seed oils extracted cold press are found as 0.28 and 0.33%, this value was determined as 0.35 and 0.41% in M. oleifera and M. peregrina oils obtained with Soxhlet extraction system, respectively. Re-fractive index values of both plant oils were found partly similar. Gener-ally, while saponification values of M. oleifera and M. peregrina oils obtained with cold press were determined partly high, density values of M. oleifera and M. peregrina extracted with cold press were deter-mined low. Increasing of peroxide values of M. oleifera and M. peregrina seed oils obtained with Soxhlet system can be attributed to the solvent used, applied heat and the presence of oxygen in the system. In addition, increasing of unsaponifiable matter of both oils extracted with Soxhlet extraction can be probably due to transferred into oil of more matter from seeds by solvent used. In previous study,Lalas and Tsaknis (2002)reported that density, refractive index, free fatty acids, saponi fi-cation value and Iodine values of M. oleifera oil obtained by cold press were determined as 0.899 mg/ml, 1.460 nD40 °C, 1.94%, 199.32 mg KOH/g and gI2/100 g, respectively. In previous study, M. oleifera and M.

peregrina oils extracted by Soxhlet extraction had low acid value. This can be attributed to the low action of lypolytic enzymes (Sengupta and Gupta, 1970). The lower iodine value can be caused by the lower number of peroxides due to the lesser degree of unsaturation. In other study,Anwar and Rashid, 2007determined 65.58–69.45 gI2/100 g

Io-dine value, 1.4549–1.4608 (40 o

C) refractive index, 0.8809– 0.9090 g/cm3_{density, 178.11}_{–186.67 mg KOH/g saponiﬁcation value,}

0.74–0.90% unsaponiﬁcation matter and 0.40–1.12% acidity in several Moringa oleifera oils. These results showed partly differences compared to results of M. oleifera oil from Kenya (Tsaknis et al., 1999), M. oleifera (Pakistan) (Anwar and Rashid (2007)and M. oleifera (Periyakulam-1)

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from India (Lalas and Tsaknis, 2002). The values of acidity (0.1 g and 0.17 to 0.14 and 0.11% for M. oleifera and M. peregrina obtained cold press and Soxhlet extraction) were considerably lower than M. oleifera oil from Pakistan (Anwar and Rashid, 2007).

The fatty acid pro_{ﬁle of oil samples from two Moringa cultivars} ob-tained using either cold pressing method or Soxhlet system is presented inTable 2. The oleic acid contents of M. oleifera and M. peregrina oils ex-tracted using cold pressing and Soxhlet method were determined as 75.49, 74.62% and 73.83, 71.67%, respectively. M. oleifera oil obtained using cold pressing showed the highest oleic acid content (75.49%), with M. peregrina oil showing the lowest content (71.67%). In addition, palmitic acid contents of M. oleifera and M. peregrina oils obtained using cold pressing and Soxhlet systems were 9.58 and 11.74% to 12.27 and 12.84%, respectively. Also, stearic acid contents of M. oleifera and M. peregrina oils obtained using cold pressing and Soxhlet techniques were between 2.81 and 4.76% to 3.67 and 3.92%, respectively. While stearic acid contents of M. oleifera and M. peregrina are found similar, stearic acid contents of M. oleifera and M. peregrina oils recovered using cold press and Soxhlet extraction methods showed statistically (pb .05) differences. Certain differences in fatty acid contents and com-position were observed in relation to Moringa species and the tech-niques used for recovering oil. Oleic acid was observed in oils obtained either using cold pressing or Soxhlet extraction system. Signiﬁcant dif-ferences (pb .05) were observed in terms of two Moringa species and extraction method used for producing oil. In previous study, the Moringa oleifera oil of cold press contained 0.13% lauric, 6.34% myristic, 5.70% stearic, 71.60% oleic, 0.77% linoleic, 0.20% linoleinic, 3.52% arachidic, 2.24% eicosenoic and 6.21% behenic (Lalas and Tsaknis, 2002). Ashraf and Gilani (2007)reported that M. oleifera seed oil contained 2.09% stearic, 1.27% linoleic, 1.75% linolenic, 74.99% oleic acids. In other study,Anwar and Rashid (2007)determined 5.51_– 6.50% palmitic, 0.97–1.46% palmitoleic, 4.14–5.88% stearic, 67.79– 76.00%oleic, 0.65–1.29% linoleic, 0.18–0.30% linolenic, 2.76–4.08% C20:0, 1.20–2.60% C20:1 and 5.00–6.81% C22:0 in M. oleifera oil. Appre-ciable variations were observed from M. oleifera oils obtained from

originating from Malawi (Tsaknis et al., 1998), Pakistan (Anwar and Bhanger, 2003) and India (Lalas and Tsaknis, 2002). The fatty acid con-tents in M. oleifera and M. peregrina oils showed considerably differ-ences reported in these studies. Signiﬁcant differences were observed among fatty acid contents of M. oleifera and M. peregrina. Moringa peregrina oils contained about 70.5% oleic acid (Tsaknis et al., 1998). The fatty acid composition of Moringa oil due to high oleic acid content was similar to that of olive (Tsaknis et al., 1998; Anwar and Rashid, 2007; Lalas and Tsaknis, 2002; Ashraf and Gilani, 2007).

The tocopherol proﬁles of M. oleifera and M. peregrina oils obtained using two different methods are presented in Table 3. The α-tocopherol contents of M. oleifera and M. peregrina oils recovered using cold pressing and solvent-based Soxhlet systems were found as 139.61 and 137.89 mg/kg to 142.74 and 151.67 mg/kg, respectively. Theα-tocopherol, ᵞ-tocopherol and δ-tocopherol contents of Moringa oils recovered using Soxhlet method were more than those in Moringa oils recovered using cold press system. Tocopherol contents of M. peregrina oil obtained using Soxhlet extraction were higher than those in M. oleifera oil. Signiﬁcant differences in tocopherol contents of M. oleifera and M. peregrina oils obtained with cold press and soxhlet ex-traction were observed (pb .05). The α-tocopherol, ᵞ-tocopherol and δ-tocopherol contents of M. oleifera and M. peregrina oils were higher than those in M. concanensis l (72.1, 9.26, 33.9 mg/kg) (Manzoor et al., 2007) and M. peregrina oil (145, 58.0 and 66.0 mg/kg) samples (Tsaknis et al., 1998). Lalas and Tsaknis (2002) observed 0.51 mg/100 g α-tocopherol, 2.54 mg/100 g ᵞ-tocopherol and 0.36 mg/100 gδ-tocopherol in M. oleifera oil obtained using cold press-ing system. In other study Morpress-inga oil contained 101.46 mg/kg α-tocopherol, 39.54 mg/kgδ-tocopherol and 75.67 mg/kg δ-tocopherol (Tsaknis et al., 1999).Anwar and Rashid (2007)reported that M. oleifera oils contained 15.38_{–140.5 mg/kg α-tocopherol, 4.47–93.70 mg/kg} ᵞ-to-copherol and 15.51–71.16 mg/kg δ-tocopherol. Generally, the levels of α-tocopherol, ᵞ-tocopherol and δ- tocopherol contents of M. oleifera and M. peregrina oils obtained using either of the extraction method were partly higher than those of M. oleifera oils shown in literatures (Tsaknis et al., 1998; Anwar and Rashid, 2007; Tsaknis et al., 1999; Lalas and Tsaknis, 2002; Abd El Baky and El-Baroty, 2013).

Sterol contents of M. oleifera and M. peregrina oils recovered using cold pressing and Soxhlet techniques are presented in Table 4.

Table 1

Physico-chemical properties of M. oleifera and M. peregrina oils.

Cold pressing Soxhlet extraction

Properties M. oleifera M. peregrina M. oleifera M. peregrina

Acid value (mg KOH/g) 0.19 ± 0.03⁎a 0.17 ± 0.02b 0.14 ± 0.05a 0.11 ± 0.03b Peroxide value (meq O2/kg) 0.18 ± 00.07a⁎⁎ 0.16 ± 0.03b 0.35 ± 0.09a 0.27 ± 0.11b

Density (g/cm3

; 25 °C) 0.819 ± 0.013b 0.828 ± 0.021a 0.847 ± 0.017a 0839 ± 0.011b Refractive Index (nD) 1.4359 ± 0.0001b 1.4371 ± 0.0002a 1.4399 ± 0.0003a 1.43860.0003b Iodine value (g I2/100g) 67.53 ± 1.37b 68.44 ± 2.17a 66.24 ± 1.58b 67.51 ± 1.19a

Saponiﬁ,cation value (mg/KOH/g) 181.3 ± 3.67 178.7 ± 2.71 177.6 ± 2.86a 176.5 ± 1.79b Unsaponiﬁcation matter (%) 0.28 ± 0.09b 0.33 ± 0.07a 0.35 ± 0.05b 0.41 ± 0.03a ⁎ Mean ± standard deviation.

⁎⁎ Values in each row with different letters are signiﬁcantly different (p b .05).

Table 2

Fatty acid composition of M. oleifera and M. peregrina oils (%). Cold pressing Soxhlet extraction Fatty acids M. oleifera M. peregrina M. oleifera M. peregrina Lauric 2.17 ± 0.28⁎a 1.93 ± 0.13b 1.74 ± 0.21a 1.63 ± 0.09b Myristic 0.78 ± 0.03a⁎⁎ 0.61 ± 0.07b 0.66 ± 0.05b 0.71 ± 0.09a Palmitic 9.58 ± 0.32b 11.74 ± 0.56a 12.27 ± 0.18b 12.84 ± 0.24a Palmitoleic 3.17 ± 0.41a 1.83 ± 0.13b 1.83 ± 0.21b 2.09 ± 0.17a Stearic 2.81 ± 0.13b 4.76 ± 0.23a 3.67 ± 0.19b 3.92 ± 0.26a Oleic 75.49 ± 0.67a 74.62 ± 0.93b 73.83 ± 0.71a 71.67 ± 0.52b Linoleic 1.69 ± 0.17b 1.84 ± 0.38a 1.27 ± 0.13b 1.38 ± 0.25a Linolenic 1.87 ± 0.21b 1.97 ± 0.16a 1.73 ± 0.09a 1.62 ± 0.11b Arachidic 5.71 ± 0.23a 3.84 ± 0.16b 2.87 ± 0.27a 1.69 ± 0.07b Eicosenoic 1.43 ± 0.09a 1.13 ± 0.07b 1.71 ± 0.11b 1.86 ± 0.09a Behenic 5.83 ± 0.13a 2.74 ± 0.18b 4.42 ± 0.46a 3.57 ± 0.32b ⁎ Mean ± standard deviation.

⁎⁎ Values in each row with different letters are signiﬁcantly different (p b .05).

Table 3

Tocopherol contents of M. oleifera and M. peregrina oils (mg/kg). Cold pressing Soxhlet extraction Tocopherols M. oleifera M. peregrina M. oleifera M. peregrina α-tocopherol 139.61 ± 1.46⁎a 137.89 ± 2.83b 142.74 ± 2.64b 151.67 ± 1.89a ᵞ- tocopherol 65.73 ± 1.54a⁎⁎ 63.68 ± 0.87b 69.84 ± 0.56b 71.51 ± 1.06a δ-tocopherol 62.51 ± 1.13a 61.49 ± 0.76b 65.27 ± 0.58a 63.58 ± 0.97b ⁎ Mean ± standard deviation.

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Campesterol, stigmasterol,β-sitosterol and δ5-avenasterols were the most predominant of the sterols detected in M. oleifera and M. peregrina oils extracted using either extraction system. Stigmasterol contents of M. oleifera and M. peregrina oils recovered using cold pressing and Soxhlet methods were found as 19.62 and 17.84% to 18.54 and 15.41%, respectively. Similarly, β-sitosterol contents of M. oleifera and M. peregrina oils extracted with cold press change as 48.56 and 46.81%, β-sitosterol contents of M. oleifera and M. peregrina oils obtained by Soxhlet extraction system were determined as 47.56 and 45.84%, re-spectively.Lalas and Tsaknis (2002)observed that cold pressed M. oleifera oil contained 15.81% campesterol, 23.10% stigmasterol, 45.58% β-sitosterol and 18.46% δ5-avenasterol. Seed oil obtained from wildly grown M. oleifera in Pakistan contained 46.16%_{β-sitosterol, 18.80%} stig-masterol, 17.95% campesterol and 9.26%δ5-avenasterol, whereas low quantities of clerosterol, 24-methylene cholesterol,δ7-campestanol and stigmastanol were also detected (Anwar and Rashid, 2007). The contents of sterols in the present analysis of Moringa oils were rather comparable with the values of M. oleifera and M. peregrina reported from several literatures (Tsaknis et al., 1999; Lalas and Tsaknis, 2002; Tsaknis et al., 1998; Al Juhaimi et al., 2016b). The sterol composition of M. oleifera oils generally showed differences compared to the conven-tional edible oils (Rossell, 1991). Generally, fatty acids, tocopherol con-tents and sterol concon-tents of Moringa oils recovered using cold pressing technique were higher in comparison to those obtained using Soxhlet system. The sterol contents reported here partly differed from those in literature. These variations can be probably due to differences in plant species, locations, climatic factors and extraction processing.

4. Conclusion

The oleic acid was observed as a predominant fatty acid in Moringa oils obtained from two plant species and it varied with the extraction method employed. The higher contents of fatty acid and tocopherol in oil obtained using cold pressing in comparison to the Soxhlet oil can be attributed to the fact that the former may contain more impuri-ties. The application of heat in Soxhlet system may also be detrimental to these oil constituents. Theα-tocopherol, ᵞ-tocopherol and δ- tocoph-erol contents of Moringa oils extracted using Soxhlet system were how-ever higher than those in cold pressed oils. Tocopherol contents varied with the species and the extraction method used. Campesterol, stigmas-terol,β-sitosterol and δ5-avenasterols were the most abundant compo-nents of sterol proﬁles of M. oleifera and M. peregrina oils obtained using both extraction systems. Generally, sterol contents of Moringa oils ob-tained using cold pressing technique were found higher while com-pared to Moringa oils extracted using Soxhlet system. Future research can focus on antioxidant effects of Moringa oils due to high tocopherol contents in several edible oils. The current study is useful for estimating

the difference in Moringa oil obtained from different species using dif-ferent post-harvest methods and based on theﬁndings cold pressing technique is recommendable method. The study further emphasize on the use of Moringa oil as a high quality oleic acid rich oil in food and me-dicinal formulations.

Acknowledgements

The authors would like to extend their sincere appreciation to the Deanship of Scientiﬁc Research at King Saud University for its funding this work through research group no. RG-1439-016.

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Table 4

Sterol contents of M. oleifera and M. peregrina oils (%).

Cold pressing Soxhlet extraction

Sterols M. oleifera M. peregrina M. oleifera M. peregrina

24-methylene cholesterol 0.97 ± 0.07⁎a 0.88 ± 0.03b 0.77 ± 0.05a 0.69 ± 0.01b

Campesterol 18.21 ± 0.18a⁎⁎ 15.57 ± 0.23b 17.43 ± 0.32a 13.21 ± 0.56b

Campestanol 0.61 ± 0.03a 0.54 ± 0.07b 0.39 ± 0.09a 0.27 ± 0.03b

δ7-campestanol 0.65 ± 0.05b 0.71 ± 0.03a 0.61 ± 0.07a 0.59 ± 0.11b

Stigmasterol 19.62 ± 0.34a 17.84 ± 0.56b 18.54 ± 1.03a 15.41 ± 1.17b

Ergostadienol 0.41 ± 0.01a 0.37 ± 0.03b 0.37 ± 0.09a 0.31 ± 0.03b

Clerosterol 2.21 ± 0.17b 2.86 ± 0.32a 1.95 ± 0.09a 1.83 ± 0.13b

Stigmastanol 0.78 ± 0.23b 0.81 ± 0.09a 0.59 ± 0.03b 0.63 ± 0.05a

β-sitosterol 48.56 ± 0.56a 46.81 ± 0.41b 47.56 ± 0.98a 45.84 ± 0.84b

δ7-avenasterol 0.97 ± 0.03b 1.04 ± 0.07a 0.77 ± 0.05b 0.83 ± 0.09a

δ5-avenasterol 12.74 ± 0.21a 9.71 ± 0.34b 10.38 ± 0.28a 8.83 ± 0.17b

28-isoavenasterol 0.74 ± 0.09b 0.93 ± 0.07a 0.51 ± 0.05b 0.67 ± 0.03a

δ7,14-stigmastanol 0.82 ± 0.11b 0.95 ± 0.13a 0.73 ± 0.09a 0.51 ± 0.07b ⁎ Mean ± standard deviation.

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