Rosemary Infusions: Comparison
View Latest Version
Please note this is a comparison between Version 2 by Vivi Li and Version 1 by Juliana Alexandra Barreto Peixoto.

Rosemary (Rosmarinus officinalis L., Lamiaceae family) is quite appreciated in worldwide gastronomy due to its aromatic flavor and preservative properties. In addition, this plant is also traditionally consumed in the infusion form aiming at the treatment and prevention of several health disorders, such as dyspepsia, mild spasmodic disorders, headache, depression, anxiety, respiratory disorders, minor peripheral circulatory disorders, and minor muscular and articular pain.

  • Rosmarinus officinalis L.
  • bioactive compounds
  • antioxidants
  • amino acids
  • UHPLC-ESI-QTOF-MS
  • RP-HPLC-FLD

1. Introduction

Rosemary (Rosmarinus officinalis L., Lamiaceae family) is quite appreciated in worldwide gastronomy due to its aromatic flavor and preservative properties [1,2,3,4][1][2][3][4]. In addition, this plant is also traditionally consumed in the infusion form aiming at the treatment and prevention of several health disorders, such as dyspepsia, mild spasmodic disorders, headache, depression, anxiety, respiratory disorders, minor peripheral circulatory disorders, and minor muscular and articular pain [1,3,5,6,7,8][1][3][5][6][7][8]. Most of these properties and, in particular, the antioxidant ones, are mainly attributed to the phenolic acids, phenolic diterpenes, and flavonoids present on rosemary [2,3,4,9,10][2][3][4][9][10]. Actually, the antioxidant activity is on the basis of the numerous properties attributed to these phenolic compounds, as they can act by different mechanisms of action, namely by free radical scavenging, hydrogen donation, metal ion chelation, by stimulating the cell antioxidant defenses, among others [11,12,13][11][12][13]. This versatility of mechanisms supports the increasing number of studies demonstrating that their regular consumption, in the necessary amounts, may help to prevent the generation of reactive oxygen species (ROS) that can cause pathological oxidative damages to cells and, consequently, contribute to the development of chronic diseases [11,12,14][11][12][14].

Beyond phenolic compounds, plants are also endowed by many other antioxidants that contribute to the biological properties of plants and increase the activity of phenolics through possible synergisms [3,4,15][3][4][15]. For example, some amino acids, besides playing a key role in many physiological functions (e.g., constituents of proteins and peptides, precursors of important metabolites, nitrogen storage molecules, regulators of important metabolic pathways, etc.), can also act as efficient antioxidants [15,16,17][15][16][17]. For example, the aromatic (tryptophan, phenylalanine, and tyrosine) and the acidic (aspartate and glutamate) ones, as well as asparagine, cysteine, alanine, isoleucine, methionine and proline can help to decrease the oxidative stress by several mechanisms, namely by scavenging free radicals, reduction of hydroperoxides, chelation of pro-oxidative transition metals, or even by acting as oxidative stress biomarkers [15,16,18,19][15][16][18][19]. Moreover, amino acids are precursors and constituents of important antioxidants peptides (e.g., glutathione) and enzymes (e.g., glutathione peroxidase, glutathione reductase, catalase, and superoxide dismutase) and particularly in plants they are essential for the biosynthesis of important bioactive compounds, such as phenolics and glucosinolates [15,16,18][15][16][18]. Therefore, the amino acid composition of plants should also be an important object of study, since this can add even more value to plants like rosemary with bioactive properties, opening the door to novel studies.

Although there are numerous studies on the bioactive properties of rosemary, many of them intend to characterize the plant itself or to evaluate/compare extraction techniques [2,9,10[2][9][10][20][21],20,21], and not so much to characterize the way the plant is often consumed by population—the infusion form. Furthermore, there is a lack of knowledge about the free amino acid composition of rosemary, particularly in the form of infusion, as well as its impact on the antioxidant properties of this beverage.

2. Antioxidant Activity and Total Contents in Phenolics and Flavonoids

The results obtained for the antioxidant activity and total phenolic contents of the different rosemary infusions studied are presented in Table 21. The phenolic contents are in accordance with those described in previous studies on rosemary [26,27,28][22][23][24]. However, rosemary infusions seem to possess lower contents of phenolic compounds and lower antioxidant potential than some herbal infusions commonly consumed worldwide, such as green tea, chamomile, sage, lemon balm, thyme, lemongrass, and echinacea infusions [26,27][22][23].

Table 21. Antioxidant activity, total phenolics, and flavonoids of rosemary infusions from different brands.
Commercial Brand Antioxidant Activity Bioactive Compounds
DPPH Inhibition

(μg TE/mL)		 FRAP

(μg FSE/mL)		 ORAC

(μg TE/mL)		 Total Phenolic Content

(μg GAE/mL)		 Total Flavonoids Content

(μg CE/mL)
A 16.40 ± 0.47 b 729.67 ± 55.93 a 157.51 ± 14.61 a 34.65 ± 1.71 a 31.22 ± 1.09 a
B 12.98 ± 1.53 b 564.67 ± 55.08 b 128.17 ± 12.47 b 27.65 ± 1.27 b 26.64 ± 0.52 b
C 16.65 ± 1.06 b 514.67 ± 53.93 b 91.72 ± 5.75 c 26.35 ± 1.23 bc 24.69 ± 1.93 bc
D 22.82 ± 1.77 a 451.33 ± 17.56 b 126.47 ± 11.28 b 23.06 ± 0.59 c 21.36 ± 1.29 c
The results are expressed as mean ± standard deviation. Within each column, different letters denote significant differences between mean values (p < 0.05). DPPH•, 1,1-diphenyl-2-picrylhydrazyl radical; FRAP, Ferric reducing antioxidant power; ORAC, Oxygen radical absorbance capacity; TE, Trolox equivalent; FSE, ferrous sulfate equivalents; GAE, gallic acid equivalents; CE, catechin equivalents.

Comparing the results between samples, infusions from brand A presented the highest antioxidant activity in the FRAP assay, followed nearly by brands B, C, and D infusions. A similar trend was observed for total phenolics and flavonoids: brand A infusions presented the highest contents of phenolics, whereas brand D showed the lowest contents. In the ORAC assay, the results were partially similar to those obtained for the abovementioned assays, with brand A infusions presenting significantly higher results, while brand C ones showed the lowest values. However, in the DPPH• scavenging assay, the results showed a reversed trend, with brand D infusions exhibiting the highest radical scavenging ability. 

3. Phenolic Profiling by UHPLC-ESI-QTOF-MS

In order to estimate the influence of the chemical composition on the antioxidant activity of rosemary infusions, a comprehensive phenolic profiling analysis was carried out based on the information provided by high-resolution mass spectrometry (HRMS/MS) data, MS databases search (e.g., Metlin and HMDB) and data described in the literature. When available, reference standards were used to confirm the identification of compounds by comparing their retention times and HRMS/MS data, also supporting the tentative identification of some structurally related compounds.

A total of forty-four phenolic compounds, belonging to nine different groups, were tentatively identified in rosemary infusions: seven hydroxybenzoic acids (1–4, 6, 7, and 12), seven hydroxycinnamic acids (9–11, 13, 16, 18, and 24), one flavan-3-ol (15), one flavanone (25), thirteen flavones (17, 19–23, 26–30, 34, and 37), eleven phenolic diterpenes (31–33, 35, 36, 39–44), one hydroxybenzaldehyde (5), two coumarins (8 and 14) and one pyranochromanone (38). Table 32 summarizes the identified compounds, as well as their chromatographic characteristics and HRMS data.

Table 32. Tentative identification of compounds detected in rosemary infusions by UHPLC-ESI-QTOF-MS.
Peak Retention Time (min) [M − H]− Experimental [M − H]− Theoretical Error (ppm) MS2 Product Ions Molecular Formula Family Tentative Identification Std ** Ref.
1 2.144 315.0717 315.0722 1.46 108 (42), 109 (38), 152 (100), 153 (41) C13H16O9 Hydroxybenzoic acid Dihydroxybenzoic acid hexoside A –
2 2.260 197.0454 197.0455 0.75 123 (75), 135 (100) C9H10O5 Hydroxybenzoic acid Hydroxydimethoxybenzoic acid B [2,5,20][2][5][20]
3 2.334 167.0344 167.0350 3.49 108 (81), 121 (100), 123 (82), 137 (76) C8H8O4 Hydroxybenzoic acid Hydroxymethoxybenzoic acid (I) C [20]
4 2.467 153.0195 153.0193 −1.09 109 (100) C7H6O4 Hydroxybenzoic acid Protocatechuic acid * A [20]

Table 43 presents a quantitative approach of the tentatively identified compounds. As previously mentioned, whenever available, the identified compounds were quantified using reference standards, whereas tentatively annotated compounds were semi-quantified using reference standards with structural similarity. The results are expressed as ng of compound/mL of infusion. Table 43 also contains the accumulated amounts of the different classes and subclasses of compounds in rosemary infusions, which was quite similar between all commercial brands: phenolic diterpenes constituted the main group of compounds, followed by hydroxycinnamic acids and flavan-3-ols groups. A color code ranging from high concentration (dark color) to low concentration (light color) allow us to follow the amount of each compound in the analyzed infusions. In addition, Figure 1 presents the total ion chromatograms (TIC) of the different infusions, where the similarities between them can be noticed.

Antioxidants 10 00500 g001 550
Figure 1. Total ion chromatograms (TIC) of rosemary infusions from different brands, analyzed by UHPLC-ESI-QTOF-MS/MS.

Table 43. Quantification of compounds detected in rosemary infusions by UHPLC-ESI-QTOF-MS (ng of compound/mL of infusion).
Peak Compound Std * Concentration (ng/mL of Infusion)
A B C D
1 Dihydroxybenzoic acid hexoside A 343.3 ± 1.8 ab 223.0 ± 23.5 b 364.3 ± 64.9 ab 456.1 ± 5.4 a
2 Hydroxydimethoxybenzoic acid B 516.4 ± 198.5 b 1269.9 ± 32.4 a 1190.9 ± 6.3 a 1134.3 ± 35.0 a
3 Hydroxymethoxybenzoic acid (I) C 489.2 ± 82.6 a 487.1 ± 89.8 a 524.6 ± 79.5 a 544.9 ± 47.5 a
5 3.141 137.0239
4 Protocatechuic acid A 325.7 ± 46.8 a 458.5 ± 91.5 a 325.9 ± 54.8 a 456.3 ± 36.3 a 137.0244 3.82 108 (48), 119 (9), 136 (53), 137 (100) C7H6O3 Hydroxybenzaldehyde Dihydroxybenzaldehyde A –
5 Dihydroxybenzaldehyde A 1125.6 ± 37.2 a 851.8 ± 88.9 a 452.7 ± 76.4 b 436.5 ± 12.7 b 6 3.181 299.0777 299.0772 −1.53 93 (100), 137 (99) C13H16O8 Hydroxybenzoic acid Hydroxybenzoic acid-O-hexoside D [20
6]
Hydroxybenzoic acid-O-hexoside D 62.3 ± 1.2 a 33.8 ± 5.3 a 51.3 ± 8.3 a 59.4 ± 2.3 a 7 3.221 137.0246 137.0244 −1.32 93 (37) C7H6O3 Hydroxybenzoic acid 4-hydroxybenzoic acid * D
7 4-hydroxybenzoic acid D 228.8 ± 6.4 a 165.9 ± 18.4 a 216.2 ± 50.5 a[20]
257.9 ± 7.1 a 8 3.411 339.0719 339.0722 0.76 177 (100), 221 (10) C15H16O9 Coumarin Dihydroxycoumarin hexoside (I) E –
8 Dihydroxycoumarin glucoside (I) E n.d. n.d. 29.7 ± 4.9 n.d. 9 3.567 341.0876 341.0878 0.66 135 (61), 161 (20), 179 (100), 221 (25), 281 (30) C15H18O9 Hydroxycinnamic acid Caffeic acid-O-hexoside F [20]
10 3.574 325.0926 325.0929 1.00 119 (49), 163 (100) a 63.8 ± 4.0 C15H18O8 Hydroxycinnamic acid Coumaric acid hexoside E –
a 88.3 ± 10.1 a 89.5 ± 6.9 a 11 3.667 353.0878 353.0878 0.02 173 (44), 179 (69), 191 (100) C16H18O9 Hydroxycinnamic acid Caffeoylquinic acid * G [5,20][5][20]
12 3.811 167.0350 167.0350 −0.10 108 (100), 152 (55) C8H8O4 Hydroxybenzoic acid Hydroxymethoxybenzoic acid (II) C [20]
13 3.874 179.0351 179.0350 −0.65 135 (100) C9H8O4 Hydroxycinnamic acid Caffeic acid * F [1,5,20][1][5][20]
14 3.907 339.0724 339.0722

− and [M − H − C3H4O4]−, whereas compound 30 could be identified as luteolin-3′-O-(3″-O-acetyl)-β-d-glucuronide or as luteolin-3′-O-(4″-O-acetyl)-β-d-glucuronide based on fragment ions at m/z 285 and 443, which are common to both isomers, and corresponding to acetylglucuronide and acetyl moieties, respectively [2]. Notwithstanding, their unambiguous identity cannot be confirmed by the analytical method employed in this work, and this is why they were generically identified as tetrahydroxyflavone-O-acetylglucuronide isomers. In turn, compounds 34 and 37 were identified as methoxyflavones’ derivatives, due to the characteristic neutral loss of methyl groups (15 Da), and, according to data reported for rosemary [2[2][5][9][20],5,9,20], they were assigned as cirsimaritin and genkwanin. Finally, compound 25, the only flavanone identified, was assigned as hesperidin, as deducted from the loss of a rutinoside moiety, as well as based on data found in the literature search [2,5[2][5][9],9], and its identity was then confirmed with the reference standard of the compound.

The phenolic diterpenes identified in this work represent the most abundant group detected in these infusions, in accordance with other studies about rosemary phytochemical composition [2,9,21][2][9][21]. A total of eleven phenolic diterpenes have been detected, two of which (compound 33 and 41) stood out particularly due to their high concentration. Compound 33, together with compounds 31 and 35, were identified as (epi)(iso)rosmanol. These three isomeric forms (rosmanol, epirosmanol and episorosmanol) are equally described in the literature for rosemary [2,5,9][2][5][9] and cannot be distinguished by the analytical method employed in this work, exhibiting characteristic fragment ions at m/z 301 and 283 resulted, respectively, from the loss of a CO2 molecule ([M − H − CO2]−), followed by the loss of a water molecule ([M − H − CO2 − H2O]−). Similar neutral losses were found for compound 32, a rosmanol derivative that shows the base peak at m/z 359—an m/z equal to the deprotonated molecular ion of compound 40, which was previously described by Borrás-Linares et al. (2014) as epirosmanol methyl ether [2]. Compound 41 (carnosol), whose presence in rosemary is already well documented in several studies, show the characteristic base peak detected at m/z 285, which results from the loss of a CO2 molecule ([M − H − CO2]−). The same loss was also observed for compound 44, which was identified as carnosic acid, as well as for its isomer (compound 36). This phenolic diterpene is highly reported as the strongest antioxidant compound and one of the main compounds in rosemary [1,2,5[1][2][5][9][20][21],9,20,21], although it was found in the studied infusions at very low levels. This can be explained due to its instability in the presence of oxygen and heat, leading to degradation into other phenolic diterpenes, such as carnosol, rosmanol, epirosmanol, epiisorosmanol, and rosmadial, which were found in high amounts in the studied samples [9]. Compounds 39 and 42 presented the same deprotonated molecular ions and might correspond to different isomeric forms already described in the literature [2,9,20][2][9][20]. The fragment ions found in each compound at m/z 299 and 315 from losses of propyl and ethylene moieties, respectively, are typical fragments of rosmadial and safficinolide. Compound 43, characterized by the loss of a methyl group from the precursor ion, was identified as rosmaridiphenol as reported in literature and databases [2,9][2][9].

Finally, a compound that does not belong to any of the compound families discussed above, was identified as isocalolongic acid (compound 38)—a pyranochromanone with reported bioactive potential, previously identified in plants of the genus Calophyllum [36,37][32][33]—based on the positive match of MS/MS data in MS databases. To the best of our knowledge, this compound in rosemary is being described for the first time.

4. Free Amino Acid Profile by RP-HPLC-FLD

The free amino acid content obtained by RP-HPLC-FLD analysis of the different rosemary infusions is shown in Table 54. Seven proteinogenic amino acids were detected (asparagine, threonine, alanine, tyrosine, phenylalanine, isoleucine, and proline), and four of them were found in all commercial brands (alanine, phenylalanine, isoleucine, and proline). Brand C infusions presented the highest content of total free amino acids (presenting the seven amino acids detected), whereas brand A showed the lowest ones (presenting only the four amino acids common to all brands). Although rosemary infusions seem to be poor in free amino acids, it is important to note that, even in small quantities, its consumption may help to maintain the physiological levels of these amino acids in the organism and the corresponding homeostasis [16,38][16][34]. In particular, these small amounts might have contributed to the antioxidant properties of rosemary infusions, even because some of the amino acids present have their antioxidant activity well described in the literature. For example, Guidea et al. (2020) found that asparagine presented the highest capacity to scavenge superoxide radicals and to chelate metal ions, compared to the other amino acids [15]. In the same study, phenylalanine and isoleucine (both present in all the samples) showed significant abilities to scavenge DPPH• and superoxide radicals, but relatively low reducing power in FRAP assay [15]. In another piece of research, the amino acids with hydrophobic residues, such as alanine, isoleucine, phenylalanine, and proline (all detected in the samples), protected myoglobin from oxidation by peroxyl radicals [39][35]. Thus, these studies reveal the antioxidant potential of some amino acids and show that, depending on their side chain residues, they can exert their antioxidant activity by different mechanisms of actions (scavenging of different radicals, reducing power, and/or chelating capacity), to a greater or lesser degree [15,39][15][35].

Table 54. Free amino acids (ng/mL) of rosemary infusions from different commercial brands, detected and quantified by RP-HPLC-FLD.
Free Amino Acid Commercial Brand
A B C D
Aspartic acid n.d. n.d. n.d. n.d.
Glutamic acid n.d. n.d. n.d. n.d.

][37]. Thus, while antioxidant activity evaluated by FRAP and ORAC assays seem to be mainly due to the presence of phenolics, DPPH• scavenging capacity cannot be solely attributed to phenolics but also to other antioxidants.

Table 65. Pearson correlations obtained between the different spectrophotometric assays.
  DPPH• Inhibition FRAP ORAC Total Phenolic Content Total Flavonoids Content
DPPH• inhibition 1 −0.469 −0.116 −0.447 −0.573
FRAP −0.469 1 0.647 * 0.934 ** 0.917 **
Asparagine tr 1.28 ± 0.00 b 1.34 ± 0.00 a n.d.
ORAC −0.116 0.647 * 1 0.552 0.562 Serine tr n.d. n.d. n.d.
Total phenolic content −0.447 0.934 ** 0.552 1 0.943 ** Glutamine n.d. n.d. n.d. n.d.
Histidine n.d. n.d. n.d. n.d.
Glycine n.d. n.d. n.d. n.d.
Threonine
Total flavonoids content −0.573 0.917 ** 0.562 0.943 ** 1n.d. n.d. 1.27 ± 0.02 a 1.27 ± 0.01 a
Arginine n.d. n.d. n.d. n.d.
Alanine 1.03 ± 0.01 a 1.01 ± 0.03 ab 1.00 ± 0.01 ab 0.97 ± 0.00 b 11
Caffeoylquinic acid TyrosineG 322.2 ± 81.5 a 361.5 ± 79.7 a 841.3 ± 52.2 a 711.9 ± 148.6 a
n.d. n.d. 0.02 ± 0.00 n.d. 12 Hydroxymethoxybenzoic acid (II) C 423.9 ± 32.8 a 300.9 ± 15.0 a 448.2 ± 85.4 a 494.1 ± 24.4 a
13 Caffeic acid F 97.0 ± 23.0 b 460.5 ± 93.1 a 298.7 ± 6.8 ab 144.4 ± 12.4 b −0.71 177 (100) C15
14 Dihydroxycoumarin glucoside (II)H16O9 Coumarin EDihydroxycoumarin hexoside (II) E –
n.d. n.d. 60.6 ± 36.6 n.d. 15 4.427 305.0706 305.0667 −12.85 59 (43), 97 (100), 225 (40)
15 Gallocatechin HC15H14O7 15,352.1 ± 831.4 Flavan-3-ol aGallocatechin H [2 10,574.4 ± 1057.5 a 13,920.5 ± 1968.0 a 16,253.0 ± 577.3 a,9,20][2][9][20]
16 4.747 163.0398 163.0400 1.37 119 (100) C9H8O3 Hydroxycinnamic acid p-
16 p-Coumaric acidCoumaric acid * EE 157.1 ± 9.0 ab 151.4 ± 8.6 b[20]
125.6 ± 3.0 b 187.3 ± 5.4 a 17 5.077 463.0877 463.0882 1.09 301 (100) C
17 Hydroxyluteolin/Quercetin-O-hexoside21 I 97.7 ± 14.2 a 126.7 ± 13.3 aH 139.8 ± 1.6 a20O12 131.8 ± 9.2 Flavone/Flavonol aHydroxyluteolin/Quercetin-O-hexoside I [2,[5,220]][5][20]
18 5.371 521.1293 521.1301 1.47 135 (19), 161 (33), 179 (59), 197 (42), 323 (86), 359 (51) C24
18 Rosmarinic acid-O-hexoside J 1050.2 ± 279.2 aH26O13 Hydroxycinnamic acid Rosmarinic acid-O-hexoside 834.7 ± 200.3 J a 1047.5 ± 99.2 a[2,5] 917.1 ± 96.2 a[2][5]
19 5.521 593.1517 593.1512 −0.85 285 (65), 151 (22), 593 (100) C27H30O15
19 Luteolin-O-rutinoside K 679.4 ± 20.9 a 370.3 ± 83.3 ab 348.5 ± 66.1 Flavone bLuteolin-O-rutinoside 532.5 ± 28.8 K ab[2,5,
20 Luteolin/Scutellarein-O-glucuronide (I) I 1164.0 ± 37.9 a
Valine n.d. n.d. n.d. n.d.
Methionine n.d. n.d. n.d. n.d.
Tryptophan n.d. n.d. n.d. n.d.
Phenylalanine 0.57 ± 0.06 b 0.68 ± 0.01 b 0.89 ± 0.08 a 0.95 ± 0.03 a
Isoleucine 0.68 ± 0.02 b 0.76 ± 0.02 b 0.87 ± 0.04 a 0.95 ± 0.02 a
Leucine n.d. n.d. n.d. n.d.
Lysine n.d. n.d. n.d. n.d. 9,20][2][5][
Hydroxyproline9][20]
n.d. n.d. 20 5.567 461.0726 461.0726 −0.10 285 (100) 627.2 ± 86.8 bC21H18O12 Flavone Luteolin/Scutellarein-O-glucuronide (I) I [ 591.3 ± 123.1 b 893.9 ± 41.2 ab1,2,5,9,20][1][2][5][9][20]
21 5.641 447.0942
21447.0933 Luteolin-7-O−2.04 285 (100) -glucoside I 248.4 ± 8.7 a 243.9 ± 12.7 aC21 205.0 ± 8.7 ab 184.6 ± 3.4 b
n.d. n.d.
Proline 0.20 ± 0.01 a 0.18 ± 0.02 a 0.19 ± 0.02 a 0.17 ± 0.01 a H20O11 −1.56 299 (20), 315 (47), 477 (100) C22H22O12 Flavone Nepitrin I 22 Nepitrin I 2492.5 ± 14.8 a[2,5, 1804.6 ± 70.6 b 1765.4 ± 124.4 b9,20 2061.6 ± 40.8 b][2][5][9][20]
23 5.908
23 Apigenin-O-rutinoside577.1561 577.1563 0.32 269 (100) C27H30O14 Flavone Apigenin-O-rutinoside K [20]
24 6.188 359.0775 359.0772 −0.71 135 (7), 161 (100), 179 (12), 197 (52) C18H16O8 Hydroxycinnamic acid Rosmarinic acid * J [1,2,5,9,20]
9 Caffeic acid-O-hexoside F 189.6 ± 52.6 a 134.8 ± 14.3 a 126.5 ± 10.2 a 119.0 ± 9.9 a Flavone Luteolin-7-O-glucoside * I [20]
K 136.4 ± 1.2 a 65.0 ± 8.4 b [1][2][5][9][20]
10 Coumaric acid hexoside E 65.5 ± 0.6  22 5.898 477.1046 477.103953.4 ± 11.4 b 63.3 ± 1.7 b
24 Rosmarinic acid J 47,541.6 ± 10,449.0 a 42,038.5 ± 4795.7 a 50,352.6 ± 3949.0 a 46,825.6 ± 77.7 a 25 6.291 609.1825 609.1825 0.00 301 (100) C28H34O15 Flavanone Hesperidin * L [
252,5,9][2][5][9]
Hesperidin L 964.8 ± 58.4 a 869.2 ± 3.3 a 713.9 ± 84.5 a 859.9 ± 36.7 a 26 6.454 461.1091 461.1089 −0.35 283 (32), 297 (10), 461 (100) C22H22O11 Flavone Homoplantaginin I [
261, Homoplantaginin I 1729.9 ± 22.5 a 875.1 ± 69.8 b2,5 900.4 ± 108.0 b,9 1011.2 ± 19.5 b][1][2][5][9]
27 6.568 461.0728 461.0726 −0.53 285 (100) C21H18O12 Flavone Luteolin/Scutellarein-O-glucuronide (II) I [1,[2,5
27 Luteolin/Scutellarein-O-glucuronide (II), I 1288.3 ± 277.3 a 1204.7 ± 14.1 a19, 725.7 ± 157.2 a20] 1434.7 ± 22.5 a][2][5][9][20]
28 7.061 503.0840 503.0831 −1.75 285 (44), 399 (100) C
28 Tetrahydroxyflavone-O23H20O13 Flavone Tetrahydroxyflavone-O-acetylglucuronide (I) I [1,2,5,9][1][2][5][9]
-acetylglucuronide (I) I 301.3 ± 39.7 a 224.6 ± 9.7 a 178.8 ± 39.0 a 311.6 ± 22.1 a 29 7.314 503.0833 503.0831 −0.36 285 (100), 399 (11) C23H20O13 Flavone Tetrahydroxyflavone-O-acetylglucuronide (II) I [1
29 Tetrahydroxyflavone-O-acetylglucuronide (II), I 267.7 ± 48.0 a 253.7 ± 0.6 a2,5 162.0 ± 35.5 a,9 310.2 ± 12.1 a][1][2][5][9]
30 7.741 503.0828 503.0831 0.63 285 (100), 443 (14) C23H20O13 Flavone Tetrahydroxyflavone-O-acetylglucuronide (III) I [1
30 Tetrahydroxyflavone-O-acetylglucuronide (III), I 223.9 ± 47.5 a 244.7 ± 7.9 a2,5 139.2 ± 23.8 a,9 253.8 ± 2.4 a][1][2][5][9]
31 8.365 345.1715 345.1707 −2.18 301 (100) C20H26O5 Phenolic diterpene (Epi)(iso)Rosmanol (I) M [2,5,9,20][2][5][9][20]
31 (Epi)(iso)Rosmanol (I) M 802.7 ± 258.7 b 12,960.9 ± 1928.9 a 15,272.0 ± 922.7  32 8.571 403.1765 403.1762 −0.68 281 (34), 299 (18), 341 (15), 359 (100) C22H28O7 Phenolic diterpene Rosmanol derivative M –
a 15,975.9 ± 1377.5 a
32 Rosmanol derivative M 4941.4 ± 334.6 a 3604.9 ± 1072.5 a 6538.5 ± 1189.6 a 6249.1 ± 578.8 a 33 8.691 345.1719 345.1707 −3.34 283 (44), 301 (100)
33 (Epi)(iso)Rosmanol (II)C M 28,364.3 ± 194.3 a20H26 21,466.6 ± 3748.7 aO5 Phenolic diterpene (Epi)(iso)Rosmanol (II) M 24,849.2 ± 98.3 a[2, 22,977.6 ± 1608.3 a5,9,20][2][5][9][20]
34 8.718 313.0716 313.0718 0.52 283 (100), 298 (83) C17H14O6 Flavone
34 Cirsimaritin N 32.1 ± 2.3 a 32.1 ± 6.1 Cirsimaritin N a 43.6 ± 5.5 a[ 41.1 ± 5.0 a2,5,20][2][5][20]
35 8.798 345.1711 345.1707 −1.02 283 (100), 301 (19)
35 (Epi)(iso)Rosmanol (III) M 13,647.7 ± 2013.1 a 2698.7 ± 267.9 C20H26O5 Phenolic diterpene (Epi)(iso)Rosmanol (III) M b 5604.5 ± 456.7 b[ 5888.5 ± 235.3 b2,5,9,20][2][5][9][20]
36 8.848 331.1918 331.1915 −0.96 283 (26), 287 (12), 331 (100) C20H28O4 Phenolic diterpene Carnosic acid isomer M [2,5
36 Carnosic acid isomer, M 1185.6 ± 55.0 a 832.9 ± 63.0 b 602.7 ± 52.3 b9,20][2][5][9][20]
810.8 ± 37.1 b 37 8.961 283.0610 283.0612 0.70 268 (100) C16H12O5 Flavone Genkwanin * O [2,5,9,20]
37 Genkwanin[ O 0.7 ± 0.2 a2][ 1.7 ± 0.8 a 1.8 ± 0.3 a5][9][20]
1.1 ± 0.2 a 38 9.051 387.1819 387.1813 −1.49 179 (8), 283 (43), 343 (100) C22H28O6
38 Isocalolongic acid M 1225.6 ± 145.0 b 9520.2 ± 1416.1 a 11,964.3 ± 511.7 a 7809.5 ± 1487.5 Pyranochromanone Isocalolongic acid M –
a 39 9.345 343.1550 343.1551 0.29 299 (34), 343 (100)
39C20H24O5 Rosmadial/Safficinolide (I)Phenolic diterpene Rosmadial/Safficinolide (I) M M[ 10,479.4 ± 2532.0 a 733.9 ± 57.8 b 1358.8 ± 208.3 b2,9,20][2][9][20]
2384.1 ± 6.5 b 40 9.445 359.1864 359.1864 0.00 283 (89), 284 (100), 285 (49), 300 (52), 317 (64) C21H28O5 Phenolic diterpene Epirosmanol methyl ether M [2]
40 Epirosmanol methyl ether M 999.0 ± 30.4 a 126.5 ± 8.6 b 278.2 ± 53.3 b 826.5 ± 0.1 a 41 9.488 329.1766 329.1758 −2.33 285 (100) C20H26O4 Phenolic diterpene Carnosol M [2,9,20][2][9][20]
42 9.591 343.1552 343.1551 −0.30 315 (55), 343 (100) C20H24O5 Phenolic diterpene Rosmadial/Safficinolide (II) M [2,9,20][2][9][20]
43 9.725
41 Carnosol M 12,227.1 ± 2277.0 c 23,022.8 ± 260.8 b 34,392.3 ± 410.0 a 27,136.5 ± 2089.0 ab
42 Rosmadial/Safficinolide (II) M 1506.4 ± 125.5 a 588.3 ± 18.2 b 1707.7 ± 200.5 a 1095.7 ± 38.7 ab315.1957 315.1966 2.76 285 (100) C20H28O3 Phenolic diterpene Rosmaridiphenol M [2,9][2][9]
43 Rosmaridiphenol M 707.7 ± 121.2 a 235.7 ± 37.9 b 546.2 ± 88.9 ab 407.9 ± 5.5 ab 44 9.838 331.1912 331.1915 0.86 287 (100) C20H28O4 Phenolic diterpene Carnosic acid *
44 Carnosic acid M 90.1 ± 38.2 M b[2,5,9 456.7 ± 38.1 a 580.4 ± 22.3 a 183.0 ± 8.7 b,20][2][5][9][20]
*, Confirmed by reference standard. **, Reference standard used for quantification or semi-quantification: A: Protocatechuic acid, B: Syringic acid, C: Vanillic acid, D: 4-hydroxybenzoic acid, E: p-Coumaric acid, F: Caffeic acid, G: Chlorogenic acid, H: Catechin, I: Luteolin-7-O-glucoside, J: Rosmarinic acid, K: Rutin, L: Hesperidin, M: Carnosic acid, N: Hesperetin, O: Genkwanin.

     Total
154,094.7
141,602.1
180,129.0
168,924.0
 
Antioxidants 10 00500 i001
     Total Phenolic acids
51,813
46,984
56,002
52,398
      
Total Hydroxybenzoic acids
2390
2939
3121
3403
      
Total Hydroxycinnamic acids
49,423
44,046
52,881
48,995
     Total Flavonoids 24,979 17,518 19,889 24,344
      Total Flavan-3-ols 15,352 10,574 13,921 16,253
      Total Flavones 8662 6074 5255 7231
      Total Flavanones 965 869 714 860
     Total Phenolic diterpenes 74,952 66,728 91,731 83,936
     Total Hydroxybenzaldehydes 1126 852 453 437
     Total Coumarins n.d. n.d. 90 n.d.
     Total Pyranochromanones 1226 9520 11,964 7810
For each commercial brand, the concentrations are expressed as mean ± standard deviation (ng/mL of infusion) of the infusions prepared in triplicate. Within each line, different letters represent significant differences between commercial brands, at p < 0.05. n.d., not detected. *, Reference standard used for quantification or semi-quantification: A: Protocatechuic acid, B: Syringic acid, C: Vanillic acid, D: 4-hydroxybenzoic acid, E: p-Coumaric acid, F: Caffeic acid, G: Chlorogenic acid, H: Catechin, I: Luteolin-7-O-glucoside, J: Rosmarinic acid, K: Rutin, L: Hesperidin, M: Carnosic acid, N: Hesperetin, O: Genkwanin.

Considering the chemical characterization of the compounds detected in rosemary infusions, their tentative identification will be discussed by classes or subclasses. In relation to hydroxybenzoic acids, most of the compounds identified are structurally related with each other: compounds 1 and 6 were tentatively identified as hexosides of compounds 4 (protocatechuic acid) and 7 (4-hydroxybenzoic acid), respectively, because of the loss of a hexose moiety (162 Da) which generated fragments ions at m/z 153 and 137, and which correspond to their respective aglycones. Moreover, the typical fragments at m/z 109 and 93, also detected in compounds 4 and 7, respectively, and originated due to loss of CO2 from the respective precursor ions (153 and 137) also helped to better elucidate the identification of compounds 1 and 6. As far as we know, compound 1 (a dihydroxybenzoic acid hexoside) is being identified for the first time in rosemary infusions. Similarly to compounds 4 and 7, compound 3 (hydroxymethoxybenzoic acid) also loses a CO2 moiety (44 Da) from the corresponding precursor ion, characteristic of hydroxybenzoic acid derivatives [20]. Moreover, it also presents a fragment ion at m/z 108, which is commonly found in MS/MS databases of hydroxymethoxybenzoic acid isomers, and which was also detected for compound 12, another hydroxymethoxybenzoic acid isomer. Compound 2 (hydroxydimethoxybenzoic acid), unlike others, generate a base peak at m/z 135 through the successive loss of a water molecule and a CO2 molecule ([M − H − H2O − CO2]−) and a m/z 123 [M − H − C3H6O2]− fragment, which seems to result from the loss of a methoxyethenol group.

The only hydroxybenzaldehyde derivative found in rosemary infusions (compound 5) was identified as dihydroxybenzaldehyde. The pattern of fragmentation was quite similar to MS spectra available on databases, which facilitated the identification of the compound. The more relevant fragment ions at m/z 108 and 119 resulted, from the loss of the aldehyde moiety and of a water molecule, respectively. As far as we know, this is the first time that dihydroxybenzaldehyde is being described for rosemary infusions.

Regarding the coumarins class, two compounds with the same deprotonated molecular ions were identified as dihydroxycoumarin hexoside I (compound 8) and dihydroxycoumarin hexoside II (compound 14). These compounds, apparently detected for the first time in rosemary infusions (only in brand C), presented the same base peak at m/z 177 as a result of the neutral loss of a hexose moiety (162). Further characterization of these compounds is required, as they may correspond to various dihydroxycoumarin hexosides, such as esculin, cichoriin, or daphnin, already found in other plants and with reported biological properties [29,30,31,32,33,34][25][26][27][28][29][30]. For example, esculin has been proved to inhibit lipid peroxidation and to scavenge hydroxyradicals, as well as to possess anti-inflammatory activity and gastroprotective effects [30,33][26][29]. Interestingly, all of these properties are also described for rosemary and, to some extent, might be associated with these compounds (even though they have been found in quite low quantities).

As observed for hydroxybenzoic acids, identified hydroxycinnamic acids are structurally related to each other. For compounds 9 and 10, the loss of a hexose moiety (162 Da) leads to fragment ions at m/z 179 and 163, together with their correspondingly characteristic fragment ions at m/z 135 and 119, both resulted from the loss of a CO2 moiety, which allowed their identification as hexosides of the compounds 13 (caffeic acid) and 16 (p-coumaric acid). As far as we know, coumaric acid hexoside (compound 10) was detected for the first time in rosemary. Compound 11 was identified as caffeoylquinic acid according to its characteristic fragments at m/z 191 (quinic acid), m/z 179 (caffeic acid), and the characteristic m/z 173 (loss of a water from quinic acid) of 4-caffeoylquinic acid (cryptochlorogenic acid) where this fragment represents the base peak, and its identity was confirmed by its reference standard. The most relevant hydroxycinnamic acid found in rosemary infusions was rosmarinic acid (compound 24). This compound, which presented the highest concentrations in all infusions representing around 28–31% of the total compounds detected, is well described in literature as a major compound of the plant [1,2,5,9][1][2][5][9] and, in fact, is considered a “family marker” for the Lamiaceae family [35][31]. It is noteworthy that rosmarinic acid is endowed by numerous beneficial properties, such as antioxidant, antiulcerogenic, antidepressant, and anti-inflammatory properties and, therefore, is considered one of the main responsible for the effects reported for rosemary infusions [3,4,5,8][3][4][5][8]. Concerning its fragmentation pattern, as a caffeic acid derivative, rosmarinic acid presented, as described, a typical fragment ion at m/z 179 corresponding to caffeic acid, as well as two caffeic acid fragments at m/z 161 and 135, resulted from the loss of a water molecule and a CO2 moiety, respectively. The fragment ion found at m/z 197 is also characteristic of rosmarinic acid and corresponds to the 2-hydroxy derivative of hydrocaffeic acid [20]. Additionally, a hexoside of rosmarinic acid was also detected in rosemary (compound 18), showing the characteristic fragments of rosmarinic acid and [M − H − C6H10O5]− and [M − H − C9H10O5]− ions—resultant from the loss of the hexose and the 2-hydroxy-hydrocaffeic acid moieties, respectively—in the MS/MS spectra.

Flavonoids represented one of the main groups of compounds found in rosemary infusions, although the results were a bit dissimilar between the three identified flavonoid subclasses. Despite being the only flavan-3-ol identified in the studied infusions, compound 15 represents around 7–10% of the total compounds detected, making the flavan-3-ol subclass the main flavonoid subclass detected. The MS/MS data of compound 15 showed fragment ions at m/z 97 and 225, which are well documented in literature and databases as gallocatechin. Gallocatechin is well described as one of the major flavonoids found in rosemary extracts [2,9,20][2][9][20].

Flavones’ subclass includes a total of thirteen compounds identified at relatively low abundance, accounting for about 3–6% of the total compounds detected in the different rosemary infusions. Compounds 17, 21, 22, and 26 were identified as hexosides of hydroxyluteolin/quercetin, luteolin, nepetin and hispidulin, respectively, because of the neutral loss of a hexose moiety (162 Da), which resulted in the respective aglycone. For compound 17, two different flavonoids (the flavone hydroxyluteolin-O-glucoside and the flavonol quercetin-O-glucoside) are documented to be present in rosemary [2,5,20][2][5][20]. Although it was not possible to completely establish the identification of compound 17 due to the lack of standards, this compound might correspond to an hexoside of the flavone hydroxyluteolin, as this compound did not contain a fragment ion at m/z 300, which is usually found for quercetin derivatives. The unambiguous identification of compound 21 was confirmed by comparing its retention time and MS/MS data with its corresponding reference standard. Compounds 19 and 23 were identified as rutinosides of luteolin and apigenin, as deducted from the neutral loss of 308 Da and the resulting fragment ions at m/z 285 and 269, respectively. Compound 19 also exhibited a characteristic fragment ion of luteolin, at m/z 151, that helped to confirm the identity of the compound.

Like hexosides and rutinosides, glucuronide compounds can be readily identified based on the neutral loss of the glucuronide moiety (176 Da), which results in the corresponding aglycones ([M − H − 176]−). Thus, compounds 20 and 27 detected in rosemary infusions were identified as glucuronides of two isomers (luteolin and scutellarein), both already described for rosemary. Compounds 28, 29, and 30 were identified as three tetrahydroxyflavone-O-acetylglucuronide isomers based on the neutral loss of an acetylglucuronide moiety (218 Da), leading to a fragment ion at m/z 285, corresponding to various possible tetrahydroxyflavone (e.g., luteolin and scutelarrein). These compounds could be assigned as luteolin-O-acetylglucuronide isomers, based on data reported in the literature [2,5,9][2][5][9]. More specifically, according to Borras-Linares et al. (2014), compounds 28 and 29 could be identified as luteolin-3′-O-(2″-O-acetyl)-β-d-glucuronide isomers I and II based on the presence of fragment ions at m/z 285 and 399 which correspond to [M − H − C8H10O7]

The results are expressed as mean ± standard deviation (infusions prepared in triplicate). Within each line, different letters symbolize significant differences between commercial brands (p < 0.05). n.d., not detected; tr, traces.

5. Impact of Phenolics and Free Amino Acids on the Antioxidant Potential

In order to estimate the impact of phenolics in the antioxidant activity of rosemary infusions, a Pearson correlation analysis between the total phenolics and flavonoids contents and DPPH• scavenging, FRAP, and ORAC assays was performed (Table 65). High correlations were found between the total phenolics and flavonoids and FRAP values, suggesting that phenolics (including the flavonoid class) of rosemary infusions exert antioxidant activity preferentially via electron transfer mechanisms. Although rosemary infusions also presented high capacity to inhibit peroxyl radicals by H atom transfer mechanisms in the ORAC assay, no significant correlations were detected between the total contents of phenolics and flavonoids and ORAC values. Therefore, it can be assumed that other antioxidant compounds present in these infusions beyond phenolics might be able to prevent lipid peroxidation in biological systems. In fact, the results obtained for DPPH• scavenging assay also suggest the presence of other types of antioxidant compounds, since negative correlations were found between DPPH• assay and total contents of phenolics and flavonoids. In addition, negative correlations were also found between DPPH• scavenging assay and FRAP and ORAC assays, which can be explained based on the mechanisms of action implicated in each of these methods. While in the DPPH• scavenging assay the radical may be neutralized by radical quenching via H atom transfer or by direct reduction via electron transfer, in the FRAP method, the mechanism of the reaction is mainly via electron transfer (the Fe3+-TPTZ complex being reduced to Fe2+-TPTZ by compounds with a redox potential <0.7 V), whereas in the ORAC assay is mainly via H atom transfer (where antioxidants present in the sample transfer H atoms to peroxyl radicals and protect fluorescein from degradation by these radicals) [40,41][36

**, Correlation is significant at the 0.01 level (2-tailed); *, Correlation is significant at the 0.05 level (2-tailed). DPPH•, 1,1-diphenyl-2-picrylhydrazyl radical; FRAP, Ferric reducing antioxidant power; ORAC, Oxygen radical absorbance capacity.

In fact, compounds such as abietane diterpenes, triterpenes, and volatile compounds (e.g., 1,8-cineole, camphor, borneol, verbenone) are well documented in the literature for rosemary, as well as their respective antioxidant properties [2,3,9,10,21,42][2][3][9][10][21][38]. More specifically, the abietane diterpenes containing two hydroxyl groups in C11 and C12, such as carnosol, rosmanol, carnosic acid, and other oxidized diterpenes were mainly responsible for the free radical scavenging activity in rosemary extracts [21]. As noted in Table 43, these compounds were present in high amounts in rosemary infusions, representing the main group of compounds found. According to this, it would be expected that the antiradical activity (DPPH•) of rosemary infusions from both commercial brands C and D was higher than the other commercial brands, due to the higher concentrations of phenolic diterpenes in these samples. However, it is worth highlighting possible interactions between the identified bioactive compounds. For example, according to Almela et al. [21], flavones interact with phenolic diterpenes in a synergistic manner. Thus, the higher accumulated concentration of flavones in commercial brand D compared to brand C might enhance the synergistic effect between phenolic diterpenes and flavones, which might explain the stronger DPPH• scavenging potential of brand D infusion. Moreover, it is important to highlight that, in the DPPH• scavenging assay, both electron and H atom transfers are involved, so it is understandable that, in complex matrices such as plant infusions—in which the compounds are present in different proportions and may exert its antioxidant activity by different mechanisms of action—interactions (e.g., synergisms and antagonisms) may occur between compounds which can lead to quite variable and unexpectable results in this assay.

Besides phenolic compounds and the other compounds above mentioned, bearing in mind the data reported in the literature about the antioxidant potential of amino acids and their derivatives, it is possible that the free amino acids detected in rosemary infusions also had an impact on their antioxidant properties. Notwithstanding, it is important to note that not all amino acids are endowed with antioxidant properties, and, besides the mechanisms of action previously mentioned in Section 3.34, amino acids can also act by others—namely by enhancing the activity of antioxidant enzymes or by acting as oxidative stress biomarkers [15,43][15][39]. Nonetheless, considering the low concentration levels of amino acids (low ng/mL) compared to the high concentration levels of phenolic compounds (high ng/mL to low μg/mL), it is evident that phenolic compounds (including flavonoids and phenolic diterpenes) were the principal responsible for the antioxidant potential of rosemary infusions.

Overall, it can be said that rosemary infusions are a good source of antioxidant compounds: a cup of 200 mL of infusion—which is the amount normally prepared and ingested by consumers according to manufacturers’ recommendations—may contain around 30 mg of phenolic compounds and a mean value of 800 ng of free amino acids. In this way, it is possible that rosemary infusions may present a positive impact on the prevention of oxidative stress-related diseases.

References

  1. Achour, M.; Saguem, S.; Sarria, B.; Bravo, L.; Mateos, R. Bioavailability and metabolism of rosemary infusion polyphenols using Caco-2 and HepG2 cell model systems. J. Sci. Food Agric. 2018, 98, 3741–3751.
  2. Borras-Linares, I.; Stojanovic, Z.; Quirantes-Pine, R.; Arraez-Roman, D.; Svarc-Gajic, J.; Fernandez-Gutierrez, A.; Segura-Carretero, A. Rosmarinus officinalis leaves as a natural source of bioactive compounds. Int. J. Mol. Sci. 2014, 15, 20585–20606.
  3. Ribeiro-Santos, R.; Carvalho-Costa, D.; Cavaleiro, C.; Costa, H.S.; Albuquerque, T.G.; Castilho, M.C.; Ramos, F.; Melo, N.R.; Sanches-Silva, A. A novel insight on an ancient aromatic plant: The rosemary (Rosmarinus officinalis L.). Trends Food Sci. Technol. 2015, 45, 355–368.
  4. Sanchez-Camargo, A.D.; Herrero, M. Rosemary (Rosmarinus officinalis) as a functional ingredient: Recent scientific evidence. Curr. Opin. Food Sci. 2017, 14, 13–19.
  5. Achour, M.; Mateos, R.; Ben Fredj, M.; Mtiraoui, A.; Bravo, L.; Saguem, S. A comprehensive characterisation of rosemary tea obtained from Rosmarinus officinalis L. collected in a sub-humid area of Tunisia. Phytochem. Anal. 2018, 29, 87–100.
  6. Costa, D.C.; Costa, H.S.; Albuquerque, T.G.; Ramos, F.; Castilho, M.C.; Sanches-Silva, A. Advances in phenolic compounds analysis of aromatic plants and their potential applications. Trends Food Sci. Technol. 2015, 45, 336–354.
  7. European Medicines Agency. Available online: (accessed on 21 December 2020).
  8. Ferlemi, A.V.; Katsikoudi, A.; Kontogianni, V.G.; Kellici, T.F.; Iatrou, G.; Lamari, F.N.; Tzakos, A.G.; Margarity, M. Rosemary tea consumption results to anxiolytic- and anti-depressant-like behavior of adult male mice and inhibits all cerebral area and liver cholinesterase activity; phytochemical investigation and in silico studies. Chem. Biol. Interact. 2015, 237, 47–57.
  9. Borras Linares, I.; Arraez-Roman, D.; Herrero, M.; Ibanez, E.; Segura-Carretero, A.; Fernandez-Gutierrez, A. Comparison of different extraction procedures for the comprehensive characterization of bioactive phenolic compounds in Rosmarinus officinalis by reversed-phase high-performance liquid chromatography with diode array detection coupled to electrospray time-of-flight mass spectrometry. J. Chromatogr. A 2011, 1218, 7682–7690.
  10. Perez-Sanchez, A.; Borras-Linares, I.; Barrajon-Catalan, E.; Arraez-Roman, D.; Gonzalez-Alvarez, I.; Ibanez, E.; Segura-Carretero, A.; Bermejo, M.; Micol, V. Evaluation of the intestinal permeability of rosemary (Rosmarinus officinalis L.) extract polyphenols and terpenoids in Caco-2 cell monolayers. PLoS ONE 2017, 12, e0172063.
  11. Barreira, J.C.M. Caracterização Biológica, Química e Nutricional de Castanea Sativa Miller e Prunus Dulcis (Miller) D.A. Webb. Ph.D. Thesis, Faculty of Pharmacy of University of Porto, Porto, Portugal, 2011.
  12. Gonçalves, S.; Gomes, D.; Costa, P.; Romano, A. The phenolic content and antioxidant activity of infusions from Mediterranean medicinal plants. Ind. Crop. Prod. 2013, 43, 465–471.
  13. Niciforovic, N.; Mihailovic, V.; Maskovic, P.; Solujic, S.; Stojkovic, A.; Pavlovic Muratspahic, D. Antioxidant activity of selected plant species; potential new sources of natural antioxidants. Food Chem. Toxicol. 2010, 48, 3125–3130.
  14. Gião, M.S.; Pereira, C.I.; Pintado, M.E.; Malcata, F.X. Effect of technological processing upon the antioxidant capacity of aromatic and medicinal plant infusions: From harvest to packaging. LWT Food Sci. Technol. 2013, 50, 320–325.
  15. Guidea, A.; Zagrean-Tuza, C.; Mot, A.C.; Sarbu, C. Comprehensive evaluation of radical scavenging, reducing power and chelating capacity of free proteinogenic amino acids using spectroscopic assays and multivariate exploratory techniques. Spectrochim. Acta A Mol. Biomol. Spectrosc. 2020, 233, 118158.
  16. Kumar, V.; Sharma, A.; Kaur, R.; Thukral, A.K.; Bhardwaj, R.; Ahmad, P. Differential distribution of amino acids in plants. Amino Acids 2017, 49, 821–869.
  17. Okumoto, S.; Funck, D.; Trovato, M.; Forlani, G. Editorial: Amino acids of the glutamate family: Functions beyond primary metabolism. Front. Plant Sci. 2016, 7, 318.
  18. Garrett, A.R.; Weagel, E.G.; Martinez, A.D.; Heaton, M.; Robison, R.A.; O’Neill, K.L. A novel method for predicting antioxidant activity based on amino acid structure. Food Chem. 2014, 158, 490–496.
  19. Zou, T.B.; He, T.P.; Li, H.B.; Tang, H.W.; Xia, E.Q. The structure-activity relationship of the antioxidant peptides from natural proteins. Molecules 2016, 21, 72.
  20. Hossain, M.B.; Rai, D.K.; Brunton, N.P.; Martin-Diana, A.B.; Barry-Ryan, C. Characterization of phenolic composition in Lamiaceae spices by LC-ESI-MS/MS. J. Agric. Food Chem. 2010, 58, 10576–10581.
  21. Almela, L.; Sanchez-Munoz, B.; Fernandez-Lopez, J.A.; Roca, M.J.; Rabe, V. Liquid chromatograpic-mass spectrometric analysis of phenolics and free radical scavenging activity of rosemary extract from different raw material. J. Chromatogr. A 2006, 1120, 221–229.
  22. Gião, M.S.; Gonzalez-Sanjose, M.L.; Rivero-Perez, M.D.; Pereira, C.I.; Pintado, M.E.; Malcata, F.X. Infusions of Portuguese medicinal plants: Dependence of final antioxidant capacity and phenol content on extraction features. J. Sci. Food Agric. 2007, 87, 2638–2647.
  23. Jimenez-Zamora, A.; Delgado-Andrade, C.; Rufian-Henares, J.A. Antioxidant capacity, total phenols and color profile during the storage of selected plants used for infusion. Food Chem. 2016, 199, 339–346.
  24. Vallverdu-Queralt, A.; Regueiro, J.; Martinez-Huelamo, M.; Rinaldi Alvarenga, J.F.; Leal, L.N.; Lamuela-Raventos, R.M. A comprehensive study on the phenolic profile of widely used culinary herbs and spices: Rosemary, thyme, oregano, cinnamon, cumin and bay. Food Chem. 2014, 154, 299–307.
  25. De Rosa, S.; Mitova, M.; Handjieva, N.; Calis, I. Coumarin glucosides from Cruciata taurica. Phytochemistry 2002, 59, 447–450.
  26. Li, W.; Wang, Y.; Wang, X.; Zhang, H.; He, Z.; Zhi, W.; Liu, F.; Niu, X. Gastroprotective effect of esculin on ethanol-induced gastric lesion in mice. Fundam. Clin. Pharmacol. 2017, 31, 174–184.
  27. Parejo, I.; Jauregui, O.; Sanchez-Rabaneda, F.; Viladomat, F.; Bastida, J.; Codina, C. Separation and characterization of phenolic compounds in fennel (Foeniculum vulgare) using liquid chromatography-negative electrospray ionization tandem mass spectrometry. J. Agric. Food Chem. 2004, 52, 3679–3687.
  28. Petrulova-Poracka, V.; Repcak, M.; Vilkova, M.; Imrich, J. Coumarins of Matricaria chamomilla L.: Aglycones and glycosides. Food Chem. 2013, 141, 54–59.
  29. Tian, X.; Peng, Z.; Luo, S.; Zhang, S.; Li, B.; Zhou, C.; Fan, H. Aesculin protects against DSS-Induced colitis though activating PPARγ and inhibiting NF-κB pathway. Eur. J. Pharmacol. 2019, 857, 172453.
  30. Zurn, M.; Toth, G.; Kraszni, M.; Solyomvary, A.; Mucsi, Z.; Deme, R.; Rozsa, B.; Fodor, B.; Molnar-Perl, I.; Horvati, K.; et al. Galls of European Fraxinus trees as new and abundant sources of valuable phenylethanoid and coumarin glycosides. Ind. Crop. Prod. 2019, 139, 111517.
  31. Napoli, E.; Siracusa, L.; Ruberto, G. New tricks for old guys: Recent developments in the chemistry, biochemistry, applications and exploitation of selected species from the Lamiaceae Family. Chem. Biodivers. 2020, 17, e1900677.
  32. Guerreiro, E.; Kunesch, G.; Polonsky, J. Chromanones de l’écorce de Calophyllum recedens. Phytochemistry 1973, 12, 185–189.
  33. Lim, C.K.; Subramaniam, H.; Say, Y.H.; Jong, V.Y.; Khaledi, H.; Chee, C.F. A new chromanone acid from the stem bark of Calophyllum teysmannii. Nat. Prod. Res. 2015, 29, 1970–1977.
  34. Wu, G. Amino acids: Metabolism, functions, and nutrition. Amino Acids 2009, 37, 1–17.
  35. Matsui, R.; Honda, R.; Kanome, M.; Hagiwara, A.; Matsuda, Y.; Togitani, T.; Ikemoto, N.; Terashima, M. Designing antioxidant peptides based on the antioxidant properties of the amino acid side-chains. Food Chem. 2018, 245, 750–755.
  36. Costa, A.S.G.; Alves, R.C.; Vinha, A.F.; Barreira, S.V.P.; Nunes, M.A.; Cunha, L.M.; Oliveira, M.B.P.P. Optimization of antioxidants extraction from coffee silverskin, a roasting by-product, having in view a sustainable process. Ind. Crop. Prod. 2014, 53, 350–357.
  37. Prior, R.L.; Wu, X.L.; Schaich, K. Standardized methods for the determination of antioxidant capacity and phenolics in foods and dietary supplements. J. Agric. Food Chem. 2005, 53, 4290–4302.
  38. Sanchez-Camargo, A.D.P.; Valdes, A.; Sullini, G.; Garcia-Canas, V.; Cifuentes, A.; Ibanez, E.; Herrero, M. Two-step sequential supercritical fluid extracts from rosemary with enhanced anti-proliferative activity. J. Funct. Foods 2014, 11, 293–303.
  39. Karpets, Y.V.; Kolupaev, Y.E.; Lugovaya, A.A.; Shvidenko, N.V.; Yastreb, T.O. Effects of nitrate and L-arginine on content of nitric oxide and activities of antioxidant enzymes in roots of wheat seedlings and their heat resistance. Russ. J. Plant Physiol. 2018, 65, 908–915.
More
© Text is available under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/)
ScholarVision Creations
Feedback