Pecan nut expeller cake (PNEC), a by-product of pecan oil extraction, was characterized and the effect of packaging and storage conditions was investigated. Samples were packaged under vacuum or in normal atmosphere and stored (10 or 20 ºC) for 12 months. Its composition, water and oil absorption, fatty acids profile, lipid oxidation (thiobarbituric acid reactive substances, TBARS), antioxidant activity [Diphenyl-1-(2,4,6-trinitrophenyl)hydrazyl] radical, DPPH● and [(2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid)] radical cation, ABTS●+), and microbiological quality were analyzed. PNEC showed high lipid and fiber contents (51.4 and 21.6 g/100g, respectively) with good water and oil absorption capacity. Vacuum storage at 10 ºC preserved antioxidant capacity (TBARS: malondialdehyde= 0.23 mg/kg, IC50 DPPH = 9.0 mg/mL, and IC50 ABTS = 21 mg/mL after one year storage). Extremely low mold and yeast levels were found in PNEC in low permeability vacuum packages, regardless of the storage temperature. Thus, PNEC could retain suitable quality attributes for up to 12 months using the right combination of packaging and temperature.
Pecan nuts (
Pecan oil is an edible pressed oil extracted from the nut; it is neutral in flavor, rich in unsaturated fatty acids, mainly oleic and linoleic acids (67.55 and 20.67%, respectively, Fernándes
PNEC is not used as a food ingredient so its introduction into the food production chain could be important to reduce the economic and ecologic costs of pecan oil production. However, before attempting to use it in the bakery as almond flour is employed, it is important to improve the knowledge on its physicochemical and functional properties and shelf life quality. Thus, the objectives of this work were to characterize the PNEC and to define storage conditions that would assure its microbial and physicochemical quality within a year, making it useful to employ as a food ingredient.
Recently produced PNEC (5 kg), obtained byfirst cold mechanical pressing of organic certified (OIA, Organización Internacional Agropecuaria) shelled pecan nuts (Nucana, Entre Ríos, Argentina) was processed to obtain a homogeneous sample using a commercial food processor (Universo, Rowenta, Germany, 14 cm blade) and sieved through a 350 μm mesh.
Two different permeability films and two temperatures were studied to store samples of 20–40 g/bag, 1.5 kg per condition) in dark rooms during a one-year period. The packages used were a low permeability (pO2 = 35 cm3/m2 d at 23 ºC and pH2O = 0.2–0.5 g/m2 d) vacuum package BB4L (Cryovac, Buenos Aires, Argentina), and a high permeability (pO2 = 6000–8000 cm3/m2 d at 23 ºC and pH2O = 14–17 g/m2 d) package PD-960 (Cryovac, Buenos Aires, Argentina). Temperatures of 10 ºC and 20 ºC were chosen.
Four treatments were considered, as a combination of package atmosphere and storage temperature. The nomenclature used was L-10 and L-20 for those samples stored under low permeability vacuum package at 10 and 20 ºC, respectively; in the same way, H-10 and H-20 correspond to those stored into the high permeable package at the same temperatures. At different times of storage, samples were taken to analyze possible changes in quality to assure it could be employed in baking or as a food ingredient.
Fat content was determined in triplicate by the Soxhlet method, using petroleum ether (Bp: 35–60 ºC) as extraction solvent (Marchetti
Water and oil absorptions were determined (Chakraborty,
The results were expressed as WAC = mL/g PNEC and OAC = mL/g PNEC, respectively. Four samples of different packages were analyzed each time.
At the end of storage, appropriate serial dilutions were plated with Plate Count Agar (PCA, Oxoid, Hampshire, UK) for total mesophilic aerobic count (incubated at 30 °C for 2 d) and total psychrotrophic aerobic count (incubated at 4 °C for 7 d), with Violet Red Bile Glucose Agar (Merck KGaA, Darmstadt, Germany) for Enterobacteriaceae (incubated at 37 °C for 24 h), and with de Man, Rogosa, Sharpe agar (MRS agar, Oxoid) for lactic acid bacteria (incubated at 30 °C for 2 d). Data were expressed as log colony-forming units per gram of sample (logCFU/g). Also, total coliform counts using the most probable number method were made (MPN, 46016) (AOAC
Total lipids of PNEC were extracted using a chloroform-methanol mix (2:1, v/v) according to the Folch
Fatty acid methyl ester (FAME) content was assayed by Shimadzu GC-MS model QH-2010 Ultra and Hewlett Packard’s (HP) model HP-6890. An AT-WAX (Alltech Associates, Inc.) capillary column of length 30 m and internal diameter of 0.25 x 10−3 m packed with polyethylene glycol (0.25 mm film thickness) was used. In QH-2010 Ultra, hydrogen (> 99.999%) was used as a carrier gas at a constant flow rate of 1.4 mL/min. MS interphase and ion source temperatures were 270 °C and 220 °C, respectively. The temperatures of the injector and the detector were set at 250 °C and 270 °C, respectively. The analysis was performed using full MS scan from m/z 50 to 400. A solvent delay of 3 min was used. Injection volume of 1 µL and a split ratio of 50:1 were used as part of the GC-MS analysis method. FAME profiles were obtained by comparison of retention times with standard FAME (Supelco 37 Component FAME Mix, Cat. No.18919–1 AMP, Sigma Aldrich) previously analyzed in the same conditions.
Methanol (> 99.5% purity), n-hexane and sodium sulphate were purchased from J.T. Baker (Mexico); and potassium hydroxide from Analar (UK).
From obtained FAME profiles iodine values (IV) were calculated using the following equation: IV = (% oleic acid x 0.988) + (% linoleic acid x 1.814) + (% linolenic acid x 2.737) (Martínez and Maestri,
Afterwards, % DPPH radical scavenging activity vs. extract concentration plots were employed to calculate the extract concentration required to scavenge 50% of DPPH in the assay medium, referred to as IC50. Two extracts were prepared for each treatment and storage time and measurements were done in quadruplicate.
Different concentrations of PNEC extracts (0.5 mL) were mixed with 2 mL of the ABTS●+ solution and the mixtures were shaken during 7 min. The reduction in ABTS●+ radical was determined by reading absorbance at 734 nm. The relationship between % ABTS●+ scavenged as a function of extract concentration was measured and PNEC extracts were assayed within an appropriate range of dilutions.
Finally, the extract concentration required to scavenge 50% of ABTS in the assay medium referred to as IC50 (half-maximal inhibitory concentration) was predicted. Two extracts were prepared for each treatment and storage time; measurements were done in quadruplicate.
Analysis of variance (ANOVA, SYSTAT, Inc., Evanston, IL, USA) was carried out to test the significance of independent variables. Experimental data were reported as mean values ± the corresponding standard error of the mean (SEM) when appropriate. For simultaneous pairwise comparisons, the least significance differences (LSD) test was chosen. Differences in means and F-tests were considered significant when P < 0.05.
Proximal composition of pecan nut expeller cake
Component | g/100 g |
---|---|
Lipids | 51.4 ± 0.1 |
Dietary Fiber | 13.6 ± 0.1 |
Proteins | 13.2 ± 0.01 |
Carbohydrates | 11.96 ± 0.07 |
Water | 5.66 ± 0.03 |
Ashes | 3.67 ± 0.05 |
mean value ± SEM
Fiber, proteins and carbohydrates are main components of PNEC with the possibility of establishing hydrophilic interactions since these constituents contain hydrophilic parts, such as polar or charged side chains (Jitngarmkusol
Effect of storage conditions on mold and yeast growth in pecan nut expeller cake stored: under vacuum at 10 ºC (L-10, ◼) and 20 ºC (L-20, ●), in normal atmosphere at 10 ºC (H-10, ▲) and 20 ºC (H-20, ▼). Error bars indicate SEM.
PNEC stored in diverse conditions showed no significant differences until day 120. Beyond this time, mold and yeast counts quickly rose for PNEC in high permeability film and 20 ºC and after 220 days samples had to be discarded because of high development.
In addition, extremely low mold and yeast levels were found in PNEC in low permeability vacuum package, regardless of storage temperature (10ºC or 20ºC). Thus, this seems to be the main control parameter for microbial spoilage. When a high permeable film was employed microbial levels exhibited significant differences between temperatures.
PNEC stored in high permeability film (H-10) and 10 ºC showed similar microbial quality to low permeability vacuum package at both temperatures during the first 9 months, being significantly different from day 300 on. However, three treatments showed very low log CFU/g values at the end of experiment.
At the end of storage, treatments with low mold and yeast counts (L-10, L-20, and H-10) were tested for other microorganisms. Nevertheless, total aerobic mesophilic and psychrotrophic,
Fatty acid (FA) profile and iodine value (IV) of pecan nut expeller cake at initial, middle (t = 6 months) or end (t = 12 months) in different storage conditions
Fatty Acid (g/100g total FA) | Storage time (months) | |||||||
---|---|---|---|---|---|---|---|---|
0 | 6 | 12 | ||||||
L-10 | H-10 | L-20 | H-20 | L-10 | H-10 | L-20 | ||
C16:0 | 3.91 ± 0.04c | 5.11 ± 0.3 ab | 5.07 ± 0.03b | 5.04± 0.06b | 5.56 ± 0.05a | 5.43 ± 0.2a | 5.61 ± 0.3a | 5.02 ± 0.07b |
C16:1 | 0.07 ± 0.02a | 0.03 ± 0.005a | 0.01 ± 0.003a | 0.01 ± 0.003a | 0.05± 0.03a | 0.04 ± 0.02a | 0.04 ± 0.01a | 0.04 ± 0.02a |
C17:0 | 0.03 ± 0.02a | N.D. | N.D. | N.D. | 0.06 ± 0.02a | 0.04 ± 0.01a | 0.04± 0.01a | 0.02 ± 0.01a |
C17:1 | 0.03 ± 0.006a | N.D. | N.D. | N.D. | 0.04 ± 0.01a | 0.04 ± 0.007a | 0.02 ± 0.01a | N.D. |
C18:0 | 0.1 ± 0.01d | 0.67 ± 0.04c | 0.71 ± 0.03c | 0.68 ± 0.07c | 2.22 ± 0.3b | 2.65 ± 0.4b | 4.05 ± 0.08a | 4.27 ± 0.2a |
C18:1 | 65.1 ± 0.03a | 64.0 ± 0.02b | 64.1 ± 0.04b | 64.4 ± 0.3b | 63.8 ± 0.2b | 64.2 ± 0.1b | 63.3 ± 0.08c | 64.3 ± 0.08b |
C18:2 | 28.6 ± 0.2ab | 28.1 ± 0.3b | 28.5 ± 0.3ab | 28.8 ± 0.3a | 26.1 ± 0.3c | 26.6 ± 0.3c | 25.2 ± 0.2d | 25.2 ± 0.1d |
C18:3 | 1.33 ± 0.08a | 0.83 ± 0.05b | 0.43 ± 0.02c | 0.22 ± 0.03d | N.D. | 1.12 ± 0.2a | 0.81 ± 0.03b | 0.17 ± 0.04d |
C20:0 | 0.05 ± 0.008c | 0.05± 0.02c | 0.03 ± 0.01c | 0.02 ± 0.01c | 0.29 ± 0.03a | 0.12 ± 0.02b | 0.29 ± 0.03a | 0.11 ± 0.03b |
C20:1 | 0.1 ± 0.004bc | 0.07 ± 0.002c | 0.03 ± 0.01d | 0.02 ± 0.008d | 0.32 ± 0.05a | 0.18 ± 0.03 b | 0.32 ± 0.06a | 0.14 ± 0.04b |
C22:0 | 0.24 ± 0.05a | 0.16 ± 0.04a | 0.20 ± 0.02a | 0.20 ± 0.02a | N.D. | N.D. | N.D. | N.D. |
C22:1 | 0.38 ± 0.03a | 0.36 ± 0.02a | 0.08 ± 0.04b | 0.01 ± 0.001b | N.D. | N.D. | N.D. | 0.04 ± 0.002b |
TOTAL | 99.91 ± 0.5 | 99.79 ± 0.8 | 99.54 ± 0.5 | 99.93 ± 0.8 | 98.69 ± 1.0 | 99.32 ± 1.3 | 99.62 ± 0.8 | 99.36 ± 0.6 |
SFA | 6.09 ± 0.1d | 6.2 ± 0.4d | 6.2 ± 0.09d | 6.3 ± 0.2d | 7.9 ± 0.4c | 8.7 ± 0.6b | 10.0 ± 0.4a | 9.4 ± 0.3ab |
MUFA | 63.27 ± 0.09b | 64.1 ± 0.05ab | 64.1 ± 0.09a | 64.4 ± 0.3a | 64.2 ± 0.3a | 64.4 ± 0.2a | 63.7 ± 0.2b | 64.5 ± 0.1a |
PUFA | 29.93 ± 0.3a | 28.9 ± 0.4b | 28.9 ± 0.03b | 29.0 ± 0.3b | 27.1 ± 0.3c | 26.2 ± 0.5d | 26.0 ± 0.2de | 25.4 ± 0.1e |
IV | 114.0 ± 0.2a | 110.8 ± 0.2b | 110.5 ± 0.2b | 110.7 ± 0.5b | 104.7 ± 0.4c | 109.0 ± 0.3b | 104.8 ± 0.2c | 104.0 ± 0.2c |
N.D. = Not detected.
*Different superscripts within the same row for the products indicate that average values differ significantly (P < 0.05).
SFA = saturated FA; MUFA = monounsaturated FA; PUFA = polyunsaturated FA
In recent years a variety of studies have demonstrated that dietary polyunsaturated fatty acids have protective effects on metabolic syndrome diseases such as type 2 diabetes, hyperlipidemia, or cardiovascular disease (Koba and Yanagita,
As the TBARs methodology is commonly employed for monitoring secondary oxidation products, i.e., aldehydes or carbonyls, which may contribute to the off-flavor of oxidized oils (Haq
Effect of storage conditions on TBARs evolution (expressed as milligrams of malonaldehyde, MDA/ kg product) of pecan nut expeller cakestored: under vacuum at 10 ºC (L-10, ◼) and 20 ºC (L-20, ●), in normal atmosphere at 10 ºC (H-10, ▲) and 20 ºC (H-20, ▼). Error bars indicate SEM.
On the other hand, the use of a high permeability film facilitated lipid oxidation. In PNEC stored without O2 restriction and maintained at 10 ºC (H-10) TBARs remained constant up to day 225, when a sharp increase (> 1 mg/kg PNEC) was observed, indicating an oxidative spoilage which could be potentially harmful (Kofakowska,
These results showed that both temperatures (10 and 20 ºC) combined with vacuum package in a low permeability film could preserve PNEC during one year. Other authors had suggested a temperature of 10 ºC and oxygen restriction as optimal conditions for a 12 month-shelf life for shelled walnuts, similar to our results for PNEC, as secondary oxidation and rancidity flavor development were critical parameters to control (Jensen
From the FA profile analyses (
SFA are the most stable fatty acids and it is not expected to detect a change in their total content. Nevertheless, the observed C16:0 proportional increment is a consequence of MUFA and PUFA percentage reduction through peroxidation.
The FA profiles and observed changes at the end of vacuum-packaged refrigerated storage are in agreement with results obtained in TBARs assay, where an O2 restrictive barrier resulted in an effective method to preserve PNEC unsaturated FA. In addition, lowering storage temperature delayed lipid oxidation. Estimated iodine values (
Effect of storage conditions on antioxidant activity of pecan nut expellercake against DPPH●, expressed as IC50 (extract concentration required to scavenge 50% of the DPPH● in the assay medium): stored under vacuum at 10 ºC (L-10, ◼) and 20 ºC (L-20, ●), stored in more permeable film at 10 ºC (H-10, ▲) and 20 ºC (H-20, ▼). Error bars indicate SEM.
Effect of storage conditions on antioxidant activity of pecan nut expellercakeagainst ABTS●+, expressed as IC50 (extract concentration required to scavenge 50% of the ABTS●+ in the assay medium): stored under vacuum at 10 ºC (L-10, ◼) and 20 ºC (L-20, ●), stored in more permeable film at 10 ºC (H-10, ▲) and 20 ºC (H-20, ▼). Error bars indicate SEM.
Both antioxidant mechanisms studied (DDPH● and ABTS●+) reflected similar tendencies during storage. It is noteworthy that only samples maintained at 10 ºC with O2 restriction presented similar DPPH● values within 365 days of storage and a slightly increase in ABTS●+ IC50. This suggests that under these conditions the lipid oxidation process is clearly inhibited because the antioxidant potential of the sample remained almost unchanged.
On the other hand, 10 ºC with normal atmosphere (H-10), and 20 ºC with O2 restriction (L-20) are conditions that can prevent or delay lipid oxidation, as seen previously in
It has been reported that pecan nuts have high levels of tocopherols (> 200 mg/kg, Fernandes
Although both methods measure antioxidant capacity, their mechanisms are different. The ABTS●+ assay measures antioxidant capacity based on the ability of the antioxidant to scavenge the blue–green colored ABTS radical cation by electron donation, while the DPPH● method determines the hydrogen donating capacity of a molecule and does not produce oxidative chain reactions or react with free radical intermediates. In spite of these differences there was a significant correlation (P < 0.05) between both IC50 values.
Miraliakbari and Shahidi (
Considering the four treatments employed and the different shelf life aspects studied, we can conclude that H-20 treatment was a proper storage condition only for short periods of time (4–5 months) because of microbial growth and the consequent risk of aflatoxin development. Lowering the temperature (H-10) would control the microbial growth but at eighth months lipid peroxidation became a problem and product quality could not be assured. Finally, employing a more effective barrier (L-20 and L-10) would solve these issues, nevertheless L-20 seemed to have suffered some antioxidant loss, and had a less protective effect over FA than L-10, which resulted in the best treatment for assuring long shelf life.
PNEC was rich in unsaturated lipids, fiber, and ashes (minerals). Its high water and oil absorption capacities indicated that it could be used as a component in bakery products. Properly storage conditions were established to guarantee the optimum quality of PNEC for a year. Oxygen restriction inhibited mold and yeast development and lipid oxidation with consequent protection of fatty acids and antioxidants present in this product. At the end of the study (12 months), the combination of 20 ºC and O2 restriction resulted in a good storage condition, with slight levels of oxidation and loss in antioxidant capacity. In addition, the use of low temperatures (10 ºC) and vacuum packaging allowed for maintaining PNEC quality: good water and oil absorption capacities, high content of unsaturated fatty acids, controlled lipid oxidation, and preserved antioxidant capacity. Establishing the proper conditions to assure PNEC quality would contribute to its inclusion in different foods since has an important dietary fiber content and high quantity of unsaturated fatty acids, making it a good choice to enhance these components in foods such as bakery products or cereal bars.
This research was supported by Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET, Argentina) and Universidad Nacional de La Plata. In addition, the authors thank NUCANA S.A. (Entre Ríos, Argentina) for providing the PNEC used.