Recovery of spent coffee grounds oil using supercritical CO₂: Extraction optimisation and physicochemical properties of oil

Figures & data

Figure 1. Schematic representation of (a) accelerated solvent extraction (ASE) system and (b) Supercritical CO₂ extraction system (showing front and back views).

Figura 1. Representación esquemática del (a) sistema de extracción acelerada de solventes (ASE) y (b) sistema de extracción supercrítica de CO₂ (vistas frontal y posterior).

Table 1. The coded and uncoded levels of independent variables used in the response surface methodology design.

Tabla 1. Niveles codificados y no codificados de variables independientes utilizadas en el diseño de la metodología de superficie de respuesta.

Independent variable	Symbol	Level
		Low (–1)	Middle (0)	High (+1)
Temperature (°C)	X1	40	50	60
Pressure (bar)	X2	175	200	225
Time (h)	X3	1	2	3

Table 2. Box–Behnken design.

Tabla 2. Diseño de Box-Behnken.

Run	Extraction temperature (°C)	Extraction pressure (bar)	Extraction time (h)	Coded temperature variable	Coded pressure variable	Coded time variable
1	60	200	1	1	0	−1
2	40	175	2	−1	−1	0
3	50	200	2	0	0	0
4	40	200	3	−1	0	1
5	40	200	1	−1	0	−1
6	50	175	1	0	−1	−1
7	40	225	2	−1	1	0
8	50	225	1	0	1	−1
9	60	200	3	1	0	1
10	50	200	2	0	0	0
11	60	225	2	1	1	0
12	50	200	2	0	0	0
13	50	175	3	0	−1	1
14	50	225	3	0	1	1
15	60	175	2	1	−1	0

Table 3. Crude oil yield from Soxhlet extraction, accelerated solvent extraction, and supercritical CO₂ extraction.

Tabla 3. Rendimiento del aceite crudo a partir de la extracción Soxhlet, la extracción acelerada de solventes y la extracción supercrítica con CO₂.

Extraction method	Solvent	% Crude oil
Soxhlet extraction (6 h)	Propanol	13.75 ± 0.35^a
Accelerated solvent extraction (120°C, 1500 psi for 5 min)	Propanol	14.02 ± 0.31^a
Supercritical CO2 extraction (50°C, 200 bar for 2 h)	CO2	12.11 ± 1.33^a

Values with same lowercase letters in the same column are not significantly different (p> 0.05).

Los valores con las mismas letras minúsculas en la misma columna no son significativamente diferentes (p> 0.05).

Table 4. Crude oil yield, acid value, percentage free fatty acids (%FFAs), iodine value (IV), saponification value (SV), peroxide value (PV), and thiobarbituric acid reactive substances (TBARS) of extracted oil from spent coffee grounds using supercritical CO₂ extraction.

Tabla 4. Rendimiento del aceite crudo, índice de acidez, porcentaje de ácidos grasos libres (% FFA), índice de yodo (IV), valor de saponificación (SV), valor de peróxido (PV) y sustancias reactivas al ácido tiobarbitúrico (TBARS) del aceite extraído de café molido usado utilizando la extracción supercrítica con CO₂.

Run	Temp (°C)	Pressure (bar)	Time (h)	Crude oil yield (%)	Acid value (mg KOH/g oil)	FFAs (%, as oleic acid)	IV (g I2/g oil)	SV (mg KOH/g oil)	PV (mEqO2/kg oil)	TBARS (mg malondialdehyde/kg oil)
1	60	200	1	5.53 ± 0.02^i	4.32 ± 0.49^ef	2.17 ± 0.25^efg	85.13 ± 1.84^def	195.80 ± 4.23^efg	9.02 ± 0.22^c	1.25 ± 0.15^bcde
2	40	175	2	4.67 ± 0.19^j	4.19 ± 0.56^ef	2.19 ± 0.20^efg	86.74 ± 0.04^cd	198.20 ± 2.24^def	6.62 ± 0.35^e	1.20 ± 0.21^bcdef
3	50	200	2	13.20 ± 0.07^b	4.28 ± 0.26^ef	2.08 ± 0.08^fg	91.47 ± 1.25^a	210.37 ± 2.89^a	3.75 ± 0.25^g	0.80 ± 0.12^ef
4	40	200	3	4.99 ± 0.07^j	4.81 ± 0.39^ed	2.42 ± 0.19^def	88.36 ± 0.99^bc	203.23 ± 2.29^bcd	5.75 ± 0.32^f	0.79 ± 0.07^ef
5	40	200	1	4.76 ± .0.11^j	5.00 ± .0.04^cd	2.52 ± .0.02^cd	89.88 ± 2.06^ab	206.72 ± 4.74^abc	10.04 ± 0.15^b	1.55 ± .0.08^bc
6	50	175	1	5.56 ± 0.38^i	5.17 ± 0.10^cd	2.65 ± 0.07^cd	80.55 ± 0.67^g	185.26 ± 1.55^h	12.52 ± 0.01^a	2.54 ± 0.33^a
7	40	225	2	9.30 ± 0.26^f	5.92 ± 0.04^ab	2.97 ± 0.02^ab	85.90 ± 2.46^cde	190.65 ± 4.00^gh	6.51 ± 0.33^e	1.06 ± 0.14^cdef
8	50	225	1	9.83 ± 0.11^f	6.24 ± 0.16^a	3.08 ± 0.15^ab	83.61 ± 0.36^ef	192.31 ± 1.18^fg	10.59 ± 0.50^b	1.20 ± 0.46^bcdef
9	60	200	3	11.75 ± 0.07^d	4.78 ± 0.17^ed	2.40 ± 0.09^def	89.56 ± 0.55^ab	208.51 ± 9.87^ab	6.76 ± 0.28^e	1.00 ± 0.04^def
10	50	200	2	10.68 ± 0.07^e	4.01 ± 0.10^f	2.02 ± 0.05^g	91.80 ± 1.35^a	212.55 ± 3.61^a	3.36 ± 0.04^g	0.69 ± 0.09^f
11	60	225	2	12.36 ± 0.15^c	4.18 ± 0.28^ef	2.18 ± 0.04^efg	85.82 ± 1.31^cde	197.38 ± 3.01^def	7.71 ± 0.65^d	1.16 ± 0.05^cdef
12	50	200	2	12.96 ± 0.12^b	4.01 ± 0.07^f	2.02 ± 0.03^g	91.63 ± 1.79^a	210.76 ± 4.46^a	3.13 ± 0.06^g	0.75 ± 0.08^ef
13	50	175	3	10.76 ± 0.07^e	5.54 ± 0.45^bc	2.78 ± 0.23^bc	86.58 ± 1.63^cd	201.10 ± 3.37^cde	9.06 ± 0.08^c	1.15 ± 0.02^cdef
14	50	225	3	13.65 ± 0.08^a	6.39 ± 0.19^a	3.21 ± 0.09^a	82.79 ± 0.51^fg	190.43 ± 1.17^gh	9.99 ± 0.16^b	1.68 ± 0.11^b
15	60	175	2	7.78 ± 0.18^h	4.64 ± 0.09^def	2.33 ± 0.04^defg	85.33 ± 0.11^def	196.26 ± 6.63^def	8.63 ± 0.74^c	1.33 ± 0.21^bcd

Values are expressed as mean ± standard deviation (n= 3). Values with different lowercase letters in the same column are significantly different (p≤ 0.05).

Los valores se expresan como media ± desviación estándar (n = 3). Los valores con diferentes letras minúsculas en la misma columna son significativamente diferentes (p ≤ 0.05).

Table 5. Analysis of variance table of the quadratic response surface model for the responses of percentage crude oil yield and physicochemical properties of the oil.

Tabla 5. Tabla de análisis de varianza del modelo de superficie de respuesta cuadrática para las respuestas del porcentaje de rendimiento de aceite crudo y propiedades fisicoquímicas del aceite.

Independent variable	DF^a	Seq.-SS^b	Adj.-SS^c	Adj.-MS^d	F-value	p-value
For % crude oil yield
Regression	6	151.967	151.967	25.378	32.61	0.000
Linear	3	86.890	73.037	24.346	31.34	0.000
Pressure	1	33.504	33.504	33.504	43.13	0.000
Time	1	29.915	0.613	0.613	0.79	0.400
Temp	1	23.472	39.366	39.366	50.68	0.000
Square	2	56.099	56.099	28.040	36.11	0.000
Time×Time	1	11.859	15.288	15.288	19.68	0.002
Temp×Temp	1	44.240	44.240	44.240	56.96	0.000
Interaction	1	8.978	8.978	8.978	11.56	0.009
Time×Temp	1	8.978	8.978	8.978	11.56	0.009
Residual error	8	6.214	6.214	0.777
Lack-of-fit	6	2.332	2.322	0.389	0.20	0.947
Pure error	2	3.882	3.882	1.941
Total	14	158.181
R^2	96.07%
Adj.-R^2	93.13%
For acid value
Regression	8	8.791	8.791	1.099	40.97	0.000
Linear	3	1.849	5.094	1.698	63.30	0.000
Pressure	1	1.268	1.526	1.526	56.91	0.000
Time	1	0.077	1.637	1.637	61.02	0.000
Temp	1	0.505	0.859	0.859	32.01	0.001
Square	3	5.634	5.634	1.878	70.02	0.000
Pressure×Pressure	1	2.528	2.781	2.781	103.67	0.000
Time×Time	1	2.900	2.766	2.766	103.12	0.000
Temp×Temp	1	0.206	0.206	0.206	7.68	0.032
Interaction	2	1.307	1.307	0.654	24.37	0.001
Pressure×Temp	1	1.199	1.199	1.199	44.70	0.001
Time×Temp	1	0.108	0.108	0.108	4.03	0.091
Residual error	6	0.161	0.161	0.027
Lack-of-fit	4	0.115	0.115	0.029	1.25	0.489
Pure error	2	0.046	0.046	0.023
Total	14	8.951
R^2	98.20%
Adj.-R^2	95.81%
For percentage free fatty acids
Regression	8	2.127	2.127	0.266	73.01	0.000
Linear	3	0.425	1.263	0.421	115.56	0.000
Pressure	1	0.276	0.494	0.494	135.69	0.000
Time	1	0.020	0.399	0.399	109.46	0.000
Temp	1	0.129	0.136	0.136	37.29	0.001
Square	3	1.455	1.455	0.485	133.12	0.000
Pressure×Pressure	1	0.733	0.806	0.806	221.33	0.000
Time×Time	1	0.694	0.669	0.669	183.67	0.000
Temp×Temp	1	0.028	0.028	0.028	7.59	0.033
Interaction	2	0.248	0.248	0.124	33.99	0.001
Pressure×Temp	1	0.220	0.220	0.220	60.48	0.000
Time×Temp	1	0.027	0.027	0.027	7.50	0.034
Residual error	6	0.022	0.022	0.004
Lack-of-fit	4	0.019	0.019	0.005	3.59	0.229
Pure error	2	0.003	0.003	0.001
Total	14	2.149
R^2	98.98%
Adj.-R^2	97.63%
For iodine vale
Regression	7	158.762	158.762	22.680	67.28	0.000
Linear	3	11.564	140.222	46.741	138.65	0.000
Pressure	1	0.142	108.247	108.247	321.09	0.000
Time	1	8.247	13.651	13.651	40.49	0.000
Temp	1	3.175	11.535	11.535	34.22	0.001
Square	2	126.633	126.633	63.317	187.81	0.000
Pressure×Pressure	1	94.302	101.854	101.854	302.13	0.000
Time×Time	1	32.331	32.331	32.331	95.90	0.000
Interaction	2	20.565	20.565	10.282	30.50	0.000
Pressure×Time	1	11.730	11.730	11.730	34.79	0.001
Time×Temp	1	8.835	8.835	8.835	26.21	0.001
Residual error	7	2.360	2.360	0.337
Lack-of-fit	5	2.303	2.303	0.461	16.11	0.059
Pure error	2	0.057	0.057	0.029
Total	14	161.122
R^2	98.54%
Adj.-R^2	97.07%
For saponification value
Regression	8	969.995	969.995	121.249	19.52	0.001
Linear	3	79.910	788.677	262.892	42.33	0.000
Pressure	1	12.610	573.920	573.920	92.42	0.000
Time	1	67.200	58.381	58.381	9.40	0.022
Temp	1	0.090	38.127	38.127	6.14	0.048
Square	2	727.070	727.068	363.534	58.54	0.000
Pressure×Pressure	1	621.180	655.096	655.096	105.49	0.000
Time×Time	1	105.887	105.887	105.887	17.05	0.006
Interaction	3	163.020	163.020	54.340	8.75	0.013
Pressure×Time	1	78.604	78.604	78.604	12.66	0.012
Pressure×Temp	1	18.802	18.802	18.802	3.03	0.133
Time×Temp	1	65.614	65.614	65.614	10.57	0.017
Residual error	6	37.261	37.261	6.210
Lack-of-fit	4	34.555	34.555	8.639	6.39	0.140
Pure error	2	2.705	2.705	1.353
Total	14	1007.26
R^2	96.30%
Adj.-R^2	91.37%
For peroxide value
Regression	8	106.638	106.638	13.330	32.80	0.000
Linear	3	15.817	58.805	19.602	48.23	0.000
Pressure	1	0.506	42.238	42.238	103.92	0.000
Time	1	14.031	23.217	23.217	57.12	0.000
Temp	1	1.279	1.808	1.808	4.45	0.079
Square	3	87.731	87.731	29.244	71.95	0.000
Pressure×Pressure	1	33.249	40.308	40.309	99.17	0.000
Time×Time	1	52.909	54.005	54.005	132.87	0.000
Temp×Temp	1	1.573	1.573	1.573	3.87	0.097
Interaction	2	3.090	3.090	1.545	3.80	0.086
Pressure×Time	1	2.064	2.064	2.064	5.08	0.065
Time×Temp	1	1.027	1.027	1.027	2.53	0.163
Residual error	6	2.439	2.439	0.406
Lack-of-fit	4	2.240	2.240	0.560	5.64	0.156
Pure error	2	0.199	0.199	0.099
Total	14	109.077
R^2	97.76%
Adj.-R^2	94.78%
For TBARS value
Regression	7	2.972	2.972	0.425	54.05	0.000
Linear	3	0.619	2.398	0.799	101.74	0.000
Pressure	1	0.155	1.053	1.053	134.04	0.000
Time	1	0.461	1.511	1.511	192.31	0.000
Temp	1	0.003	0.050	0.050	6.34	0.040
Square	2	1.413	1.413	0.706	89.92	0.000
Pressure×Pressure	1	0.719	0.819	0.819	104.26	0.000
Time×Time	1	0.694	0.694	0.694	88.38	0.000
Interaction	2	0.940	0.940	0.470	59.85	0.000
Pressure×Time	1	0.875	0.875	0.875	111.43	0.000
Time×Temp	1	0.065	0.065	0.065	8.27	0.024
Residual error	7	0.055	0.055	0.008
Lack-of-fit	5	0.049	0.049	0.010	3.22	0.254
Pure error	2	0.006	0.006	0.003
Total	14	3.027
R^2	98.18%
Adj.-R^2	96.37%

^a DF, degree of freedom; ^b Seq-SS, sequential sums of squares; ^c Adj.-SS, adjusted sum of squares; ^d Adj.-MS, adjusted mean squares. TBARS = thiobarbituric acid reactive substances.

^a DF, grado de libertad; ^b Seq-SS, sumas secuenciales de cuadrados; ^c Adj.-SS, suma de cuadrados ajustada; ^d Adj.-MS, cuadrados medios ajustados. TBARS = sustancias reactivas al ácido tiobarbitúrico.

Table 6. Quadratic response surface model of acid value, percentage free fatty acids (%FFAs), iodine value (IV), saponification value (SV), peroxide value (PV), and thiobarbituric acid reactive substances (TBARS) of crude oil obtained from spent coffee grounds by supercritical CO₂ extraction.

Tabla 6. Modelo de superficie de respuesta cuadrática del índice de acidez, porcentaje de ácidos grasos libres (% FFA), índice de yodo (IV), valor de saponificación (SV), valor de peróxido (PV) y sustancias reactivas al ácido tiobarbitúrico (TBARS) del aceite crudo obtenido por extracción supercrítica con CO₂ de café molido usado.

Response	Quadratic polynomial model	Adj.-R^2	p-value
% Crude oil yield	= – 96.1108 + 0.0819Pressure + 2.5582Time + 3.3229Temp – 2.0288Time^2 – 0.0345Temp^2 + 0.1498Time×Temp	0.9313	0.000
Acid value	= 34.8218–0.4300Pressure-4.1864Time + 0.6161Temp + 0.0014Pressure^2 + 0.8655Time^2 – 0.0024Temp^2 – 0.0022Pressure×Temp + 0.0164Time×Temp	0.9581	0.000
%FFAs (oleic)	= 23.718–0.2447Pressure – 2.0662Time + 0.2451Temp + 0.0007Pressure^2 + 0.4257Time^2 – 0.0009Temp^2 – 0.0009Pressure×Temp + 0.0083Time×Temp	0.9763	0.000
IV	= – 265.920 + 3.483Pressure + 19.085Time – 0.360Temp – 0.008Pressure^2 – 2.950Time^2 – 0.068Pressure×Time + 0.149Time×Temp	0.9707	0.000
SV	= – 600.328 + 8.370Pressure + 39.469Time – 2.555Temp −0.021Pressure^2 – 5.339Time^2 – 0.177Pressure×Time + 0.009Pressure×Temp + 0.405Time×Temp	0.9137	0.001
PV	= 275.851–2.182Pressure – 24.901Time – 0.714Temp + 0.005Pressure^2 + 3.824Time^2 + 0.007Temp^2 + 0.0297Pressure×Time + 0.051Time×Temp	0.9478	0.000
TBARS value	= 40.776–0.3436Pressure – 6.3494Time – 0.0237Temp + 0.0008Pressure^2 + 0.4324Time^2 + 0.0187Pressure×Time +0.0127Time×Temp	0.9637	0.000

Figure 2. Response surface (3D) plots showing the effect of different extraction temperature, pressure and time on (a) crude oil yield, (b) acid value (AV), (c) percentage of free fatty acids (%FFAs), (d) iodine value (IV), (e) saponification value (SV), (f) peroxide value (PV), and (g) thiobarbituric acid reactive substances (TBARS) value of crude oil obtained using supercritical CO₂ extraction.

Figura 2. Gráficos de superficie de respuesta (3D) que muestran el efecto de diferentes temperaturas de extracción, presión y tiempo en: (a) el rendimiento de aceite crudo, (b) el índice de acidez (AV), (c) el porcentaje de ácidos grasos libres (% FFA), (d) el valor de yodo (IV), (e) el valor de saponificación (SV), (f) el valor de peróxido (PV) y (g) el valor de sustancias reactivas al ácido tiobarbitúrico (TBARS) del aceite crudo obtenido mediante la extracción supercrítica con CO₂.

Figure 3. Optimum region overlaying the curves of the responses crude oil yield and its acid value (AV), percentage free fatty acids (%FFAs), iodine value (IV), saponification value (SV), peroxide value (PV) and thiobarbituric acid reactive substances (TBARS) value as a function of the extraction temperature, extraction time, and extraction pressure used during the extraction process.

Figura 3. Superposición de la región óptima a las curvas de respuesta del rendimiento del aceite crudo y su valor ácido (AV), porcentaje de ácidos grasos libres (% FFA), valor de yodo (IV), valor de saponificación (SV), valor de peróxido (PV) y valor de sustancias reactivas al ácido tiobarbitúrico (TBARS) en función de la temperatura, el tiempo y la presión utilizadas durante el proceso de extracción.

Table 7. Fatty acid composition of crude oil from spent coffee grounds using supercritical CO₂ extraction.

Tabla 7. Composición de ácidos grasos del aceite crudo obtenido de granos de café usados utilizando la extracción supercrítica con CO₂.

Fatty acid composition (g/100 g, %)	Crude oil from spent coffee grounds			Rice bran oil	Soybean oil	Sesame oil
	Conditions of supercritical CO2 extraction
	40°C/175 bar/2 h	60°C/200 bar/3 h	50°C/200 bar/2 h
Butyric acid (C4:0)	0.20 ± 0.00^c	0.21 ± 0.01^c	3.73 ± 0.86^a	0.28 ± 0.22^c	0.14 ± 0.01^d	2.59 ± 0.24^b
Caproic acid (C6:0)	0.07 ± 0.02	ND	ND	ND	ND	0.04 ± 0.00
Myristic acid (C14:0)	0.07 ± 0.00	0.07 ± 0.00	ND	0.32 ± 0.00	0.07 ± 0.00	0.01 ± 0.00
Palmitic acid (C16:0)	41.83 ± 2.12^a	36.67 ± 0.21^b	32.98 ± 0.81^b	19.20 ± 0.50^c	10.78 ± 0.32^d	8.87 ± 0.04^e
Margaric acid (C17:0)	0.13 ± 0.00	0.11 ± 0.00	ND	0.05 ± 0.01	0.09 ± 0.01	0.04 ± 0.01
Stearic acid (C18:0)	7.78 ± 0.56^a	7.42 ± 0.12^a	7.08 ± 0.05^a	2.01 ± 0.05^d	3.76 ± 0.09^c	5.28 ± 0.02^b
Arachidic acid (C20:0)	3.35 ± 0.42^a	3.44 ± 0.10^a	3.25 ± 0.13^a	0.86 ± 0.06^b	0.37 ± 0.02^d	0.62 ± 0.03^c
Heneicosylic acid (C21:0)	0.09 ± 0.01	0.10 ± 0.01	ND	0.01 ± 0.00	0.03 ± 0.00	ND
Behenic acid (C22:0)	0.63 ± 0.08^a	0.75 ± 0.02^a	0.67 ± 0.04^a	0.29 ± 0.02^c	0.41 ± 0.08^b	0.14 ± 0.00^d
Tricosylic acid (C23:0)	0.12 ± 0.01	0.14 ± 0.00	ND	0.03 ± 0.01	0.06 ± 0.01	ND
Lignoceric acid (C24:0)	0.27 ± 0.04	0.35 ± 0.01	0.30 ± 0.02	0.43 ± 0.06	0.16 ± 0.01	ND
Palmitoleic acid (C16:1)	ND	ND	ND	0.17 ± 0.00	0.08 ± 0.00	0.11 ± 0.00
Heptadecenoic acid (cis-10) (C17:1)	ND	ND	ND	ND	0.05 ± 0.00	ND
Oleic acid (C18:1 n9c)	7.75 ± 0.52^d	8.30 ± 0.14^d	7.50 ± 0.04^d	42.20 ± 0.07^a	26.56 ± 0.04^c	36.78 ± 0.10^b
Gondoic acid (C20:1)	0.33 ± 0.03^c	0.36 ± 0.00^c	0.36 ± 0.01^c	0.64 ± 0.06^b	0.77 ± 0.04^a	0.16 ± 0.00^d
Erucic acid (C22:1 n9)	ND	ND	ND	0.04 ± 0.01	ND	ND
Nervonic acid (C24:1)	0.20 ± 0.02	0.08 ± 0.02	ND	ND	0.05 ± 0.03	ND
Linoelaidic acid (C18:2 n6t)	ND	ND	ND	0.42 ± 0.01	0.39 ± 0.04	ND
Linoleic acid (C18:2 n6c)	34.99 ± 2.31^d	40.08 ± 0.27^c	42.45 ± 0.12^c	31.49 ± 0.07^e	50.61 ± 0.24^a	44.96 ± 0.08^b
γ-Linolenic acid (C18:3 n6)	ND	ND	ND	0.13 ± 0.01	0.52 ± 0.01	ND
α-Linolenic acid (C18:3 n3)	0.73 ± 0.05^e	0.94 ± 0.00^c	1.15 ± 0.02^b	0.88 ± 0.01^d	4.92 ± 0.01^a	0.30 ± 0.01^f
Docosadienoic acid (C22:2)	ND	ND	ND	ND	ND	0.09 ± 0.09
Eicosapentaenoic acid (C20:5 n3)	0.12 ± 0.01	0.13 ± 0.00	ND	ND	ND	ND
Saturated fatty acids (SFAs)	54.54 ± 3.27^a	49.27 ± 0.07^b	48.00 ± 0.29^b	23.45 ± 0.55^c	15.84 ± 0.11^e	17.60 ± 0.20^d
Monounsaturated fatty acids	8.28 ± 0.56^d	8.86 ± 0.12^d	7.86 ± 0.03^e	43.05 ± 0.13^a	27.51 ± 0.05^c	37.05 ± 0.10^b
Polyunsaturated fatty acids (PUFAs)	35.83 ± 2.36^e	41.15 ± 0.28^d	43.60 ± 0.10^c	32.94 ± 0.01^f	56.48 ± 0.18^a	45.35 ± 0.09^b
Total fatty acids	98.65 ± 0.35^ns	99.27 ± 0.47 ^ns	99.47 ± 0.16 ^ns	99.44 ± 0.43 ^ns	99.82 ± 0.24 ^ns	99.99 ± 0.01 ^ns
PUFA/SFA ratio (P/S index)	0.66	0.84	0.91	1.40	3.57	2.58

Data are expressed as mean ± standard deviation (n = 3). Values in each row having different lowercase letters indicate significant differences at p ≤ 0.05 (Duncan’s test); ns = not significant; ND = not detected.

Los datos se expresan como media ± desviación estándar (n = 3). Los valores de cada fila que tienen letras minúsculas diferentes indican diferencias significativas a p ≤ 0.05 (prueba de Duncan); ns = No significativo; ND = No detectado.