1. Introduction
Shortbread cookies are pastry goods characterized by density and brittleness due to high quantities of fat rich in saturated fatty acids (SFA), especially myristic, palmitic, and stearic acids [
1]. However, high consumption of SFA is associated with an increased risk of cardiovascular diseases. For this reason, much attention is focused on the improvement of their sensory and nutritional profiles through preparation of low-fat cookies using emulsifiers, interesterified shortenings, or some vegetable oils rich in monounsaturated fatty acids (MUFA) and polyunsaturated fatty acids (PUFA) [
2,
3]. The presence of oleic, linoleic, and linolenic acids in the chemical composition of vegetable oils may have a positive effect on health and may also be conducive to oxidation of a double bound they contain and thus, cause development of off-flavor and odors unacceptable to consumers [
4]. Therefore, application of plant materials as sources of many active antioxidants instead of synthetic antioxidants can be an interesting alternative in preventing the oxidation of fat contained in shortcrust pastry goods, and also because of their beneficial effects on human health. Spices and herbs, especially from the Lamiaceae family like rosemary, thyme, and marjoram, are well-known for their antioxidant activity and as excellent sources of phenolic compounds that contribute to prevention of lipid oxidation and to improving quality and nutritional value of foods [
5,
6].
Lemon balm (
Melissa officinalis L.) and hyssop (
Hyssopus officinalis L.) are also perennial herbs belonging to the Lamiacea family, cultivated worldwide. Lemon balm is an important medicinal plant and a versatile culinary herb which can be used to flavor different products owing to its particular taste. It is very useful for treating nervous agitation and for promoting sleep, and it ameliorates functional gastrointestinal complaints [
7]. Lemon balm is also used to treat herpes, gout, sores, insect bites, skin diseases, and as an insect repellent [
8]. It can be added to salads, butters, cheeses, ice cream, jams, sauces, or cookies. Lemon balm leaves, flowers, and branches are a source of volatile oils, glycosides of alcoholic triterpene acids, and phenolic compounds such as phenolic acids and flavonoids [
7]. The content of volatile oils in lemon balm is quite low compared with other members of the Lamiacea family and because of this, the price of the volatile oil is very high in the market. The volatile oil from lemon balm may be used as an antivirus, antimicrobial, and antifungal agent, effective against a series of human cancer cell lines and mouse cell line and as an antioxidant [
9]. In turn, lemon balm aqueous and alcoholic extracts are rich in phenolic acids such as rosmarinic, caffeic, chlorogenic, and ferulic acids. Presence of these compounds in extracts may affect their antibacterial, antiviral, and antioxidant activities [
10,
11]. Lemon balm extract may be helpful in terms of preventing lipid deterioration in sausages and pork meat patties and in stabilization of long-chain fatty acids of algae oil [
12].
Hyssop is an important medicinal plant used in tea blends for cough relief, antispasmodic, stomachic, and antifungal effects [
13]. Its essential oil, rich in pinocamphore,
β-pinene, myrtenol, linalool, methyleugenol, and limonene, is used in the food, pharmaceutical, and cosmetic industries [
14,
15]. Like other aromatic plants from the Lamiaceae family, hyssop extracts contain phenolic compounds, especially chlorogenic, protocatechuic, ferulic, syringic, caffeic, and rosmarinic acids [
16]. The positive correlation between phenolic acids and flavonoids present in hyssop extract may indicate its significant antioxidant activity [
17]. Among studied plant extracts, the ethanolic extract of
Hyssopus officinalis has shown a good antioxidant activity but was characterized by the lowest total polyphenols and flavonoids content [
18]. The inhibitory activity of the hyssop extract during autooxidation of lard was also observed [
19]. Hyssop extracts also inhibited lipid oxidation and degradation of heme pigments after meat cooking and storage [
20].
Stinging nettle (
Urtica dioica L.) is a medicinal and edible plant, and fodder for animals. It is used in pharmaceuticals, cosmetics, and foodstuffs. Dried nettle leaf preparations are known to alleviate symptoms associated with allergic rhinitis and are a sustainable source of textile fiber [
21]. Nettle leaves extract is used as a diuretic, in infections of urinary tract, as an adjuvant remedy in rheumatoid arthritis, in renal calculi and gout [
22]. Nettle leaves are a good source of ascorbic acid and pro-vitamin A, essential amino acids, carbohydrates, fatty acids, carotenoids, and several minerals [
23]. Stinging nettle could be used as a good natural, low-cost green pigment source, a vegetable in juice tea, and an ingredient in many dishes. Compared to barley and wheat flour, nettle flour has a much higher level of tannin, total polyphenols and content of protein, crude fiber, ash, and calcium and has a low glycemic index [
24]. Use of natural antioxidants from water nettle extract as functional ingredients significantly reduced lipid oxidation, increased color stability during storage, and improved functionality of the final product [
24]. Moreover, it also showed antimicrobial activity against some microorganisms, antiulcer activity against ethanol-induced ulcerogenesis, and analgesic effect on acetic acid-induced stretching [
25]. The capacity of nettle extracts to improve oxidative stability of brined anchovies has been also reported [
26].
The present study was undertaken to prepare shortbread cookies by partially replacing palm fat with cold-pressed walnut oil supplemented with aqueous ethanolic extracts from lemon balm, hyssop, and nettle. Color intensity and selected sensory properties of the shortbread cookies after baking were determined. Then, fat stability extracted from the cookies during their storage at room temperature by determining the peroxide, p-anisidine values, specific UV extinctions (K value), and the capacity of fat samples containing herbal extracts to scavenge DPPH radicals were evaluated.
2. Materials and Methods
2.1. Materials and Chemicals
Aqueous ethanolic extracts from plant materials, such as dried leaves of lemon balm (
Melissa officinalis L.), hyssop (
Hyssopus officinalis L.), and nettle (
Urtica dioica L.), were used as fortification agents, were prepared in advance according to Kozłowska et al. [
27]. Ingredients for making shortbread cookies were purchased in a local market (Warsaw, Poland). These were palm fat “Akofect LT M53” (AarhusKarlshamn, Malmo, Sweden), cold-pressed walnut oil “ULEI” (PROMET-T SA, Moldova), wheat flour type 480 “Szymanowska” (Polskie Młyny, Warsaw, Poland), sugar (Pfeifer & Langen Marketing, Poznań, Poland), and eggs. All the solvents (ethanol, methanol, diethyl ether, chloroform,
n-hexane) and reagents (acetic acid, potassium hydroxide, potassium iodide, potassium chloride, sodium thiosulfate, starch soluble, phenolphthalein, anhydrous magnesium sulphate) used were analytical grade and were purchased from Avantor Performance Materials Poland (Gliwice, Poland). Butylated hydroxyanisole (BHA), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and Trolox ((±)-6-hydroxy-2,5,7,8-tetramethylchromane-2-carboxylic acid) were obtained from Sigma-Aldrich (St. Louis, MO, USA). A certified fatty acids methyl ester (FAME) reference standard mixture (37 fatty acids from C4 to C24) was from Supelco (Bellefonte, PA, USA).
2.2. Shortbread Cookies Preparation
Dry ingredients comprising twice-sieved wheat flour (300 g) and sugar (100 g) were mixed using a spoon. Then eggs (50 g), whipped at medium speed using a kitchen mixer (Multiquick, Braun, Germany), were added and the mixture was blended. Fat was added (200 g) and blended for 1 min using Braun K650 Multiquick kitchen machine food processor (Germany). Then, the dough was kneaded by hand to combine the ingredients and placed in a refrigerator to cool at 4 °C (about 1h). The dough was rolled out to a thickness of 3 mm, cut into cookie shapes (5 cm diameter), placed on an aluminum tray and baked at 160 °C for 25 min in Unox convection oven (model XBC, Vigodarzere, Italy). The shortbread cookies were cooled at room temperature, packed in plastic pouches (PEHD) (every type of the cookies in a separate bag), and stored without access of light at room temperature for 3 months. They were taken out after 0, 1, 2, and 3 months of storage and used for further studies.
10 variants of fat were used in preparation of ten formulations of the shortbread cookies. For this purpose, palm fat (110 g) was weighed into flasks and dissolved in a water bath at 40 °C. Then cold-pressed walnut oil (90 g) and aqueous ethanolic extracts from lemon balm, hyssop and nettle at 3 different levels (0.02%, 0.1%, 0.2%), and synthetic antioxidant (BHA 0.02%) were added. The fat mixture was stirred using a kitchen mixer (Multiquick, Braun, Frankfurt, Germany) for 1 min and cooled in a refrigerator at 4 °C. Control samples contained fat (palm oil and cold-pressed walnut oil) without addition of antioxidants.
2.3. Fatty Acid Analysis
The fatty acid composition of the fat samples used in the preparation of the control samples was analyzed by gas chromatography (GC) after derivatization to fatty acid methyl esters with a 2 M methanolic solution of potassium hydroxide according to ISO 12966-2:2017 standard method [
28]. A Shimadzu GC-17A (Kyoto, Japan) gas chromatograph equipped with a flame ionization detector (FID) and a BPX-70 capillary column (length 30 m, i.d. 0.22 mm, film thickness 0.25 µm; Melbourne, Australia) were used. The analysis was performed with nitrogen (1.0 mL/min) as a carrier gas at the following temperature program: 60 °C held for 1 min, after which the temperature was increased to 170 °C at a rate of 10 °C/min and from 170 to 230 °C at a rate of 3 °C/min. The temperature was kept at 230 °C for the subsequent 15 min. The injector (split ratio of 100:1) and detector temperatures were set at 225 °C and 250 °C, respectively. Individual fatty acids were identified by comparing their retention times with a certified fatty acid methyl esters mix and quantified as a percentage of the total fatty acids.
2.4. Color Measurement
The color intensity of the shortbread cookies with and without the addition of different concentrations of aqueous ethanolic plant extracts and BHA was determined after baking using a tristimulus reflectance colorimeter (Minolta CM-3600d, Konica Minolta Sensing, Inc., Tokyo, Japan). It was expressed as L*, a* and b*, where L* represents lightness of color (value 100) or blackness (value 0), a* represents red (positive value) or green (negative value), and b* defines the proportion of yellow (positive value) or blue (negative value). The final result was the arithmetic mean of 12 measurements.
2.5. Sensory Evaluation
The sensory properties of the shortbread cookies samples without and with different amounts of plant extracts and BHA such as aroma, taste and overall acceptability were evaluated by ten panel members from the Food Science and Nutrition Department. The samples (each weighing around 5 g) were served to the panelists on paper plates with water to cleanse and neutralize their palates 2 h after baking. All the samples were presented with three digit codes in a randomized order. The scores for each parameter ranged from 1 to a maximum of 5, where 1 indicated worst quality and the highest score (5) represented best quality.
2.6. Fat Extraction
Fat from ground shortbread cookies (50 g) after their baking and storage (1, 2, and 3 months) was extracted by shaking (Elpan, water bath shaker type 357, Elpin-Plus, Lubawa, Poland) at ambient temperature (21 °C) for 40 min with the use of n-hexane. After filtration and separation of the fat, the solvent was removed by evaporation under reduced pressure using a rotary evaporator (Rotavapor R-215, Büchi Labortechnik, Switzerland) at 40 °C. The fat obtained was frozen (−21 °C) until further use. Chemical analysis of the fat extracted from the cookies consisted in determining peroxide value (PV), p-anisidine value (p-AV), specific UV extinctions (K232 and K268 values), and antioxidant activity (DPPH assay).
2.7. Chemical Analysis
The peroxide and
p-anisidine values and specific UV extinctions were determined according to ISO standard methods (3960:2009, 6885:2008, 3656:2011, respectively) [
29,
30,
31].
The antioxidant activity of fat samples was determined using DPPH radicals as described by Kozłowska et al. [
32,
33]. Briefly, 50 mg of each fat sample was dissolved in 3 mL of ethyl acetate. Then, 1 mL of a sample was withdrawn and diluted with 2.75 mL of ethyl acetate and 0.25 mL of a freshly prepared DPPH solution (1 mM) was added. The samples were vigorously mixed for 10 s in a vortex and stored for 20 min at room temperature in the dark. Absorbance was measured at 515 nm using a UV/Vis spectrophotometer (Model 8500, Techcomp, HongKong). The results were expressed as Trolox equivalent antioxidant capacity using a Trolox calibration curve (µmol TEAC/g of oil).
2.8. Statistical Analysis
A one-way analysis of variance (ANOVA) and Tukey’s test were used to establish the significance of differences between the means at p < 0.05. The statistical analysis was carried out with Statgraphics plus 4.0 package (Statistical Graphics Corp., The Plains, VA, USA).