Any processing method that maintains the level of compounds known for their health benefits will be of interest to the food industries. Therefore, the effects of vacuum drying, storage and freezing on the anthocyanin content and their antioxidant properties of Iresine herbstii L. flowers were investigated. The results showed that fresh samples (AEFF) had the highest amount of total anthosyanin content (8.31 ± 0.23 mg/g dry matter, expressed as cyaniding 3-glucoside equivalents), followed by 7.17 ± 0.12 mg/g solid content, 13.72% loss of vacuum dried samples (AEDF). In comparison with fresh samples, total anthocyanins in stored samples for two weeks at 5°C (AESF) and frozen samples during 1 (AEZF 1) and 3 months (AEZF 3) of storage were significantly (P < 0.01) reduced to 6.43 ± 0.24 mg/g solid content, 22.63% loss, 5.65 ± 0.33 mg/g solid content, 32.01% loss and 4.71 ± 0.51 mg/g solid content, 43.33% loss, respectively. Anthocyanins from I. herbstii L. flowers exhibited a dose-dependent (AEFF > AEDF > AESF > AEZF 1 > AEZF 3, respectively) antioxidant activity against lipid peroxidation in a linoleic acid model system as well as strong reducing power and ferrous ion chelating abilities. Moreover, the anthocyanins extracted were found to show remarkable scavenging activity on superoxide anion radicals, hydroxyl radicals, hydrogen peroxide, nitric oxide radicals and deoxyribose degradation. Based on the results obtained, we can concluded that the Iresine herbstii L. flowers may be valuable natural antioxidant sources and are potentially applicable in both pharmacy and food industry
Antioxidants are widely used as a food additive to provide protection against oxidative degradation of foods by free radicals. In order to prolong the storage of foods, several synthetic antioxidants such as butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA) are used currently. But these substances may be inappropriate for chronic human consumption. Hence the development of alternative antioxidants from natural origin has attracted considerable attention [
All the chemicals were purchased from Sigma-Aldrich Co. (St. Louis, MO), Riedal de Haën (Germany), Fluka (Switzerland), Randox (United Kingdom) and solvents were from E. Merck (Darmstadt, Germany). All of the reagents were prepared in deionized distilled water to eliminate the contamination of metal ions.
Flowers of Iresine herbstii L. were collected from the garden of College of Education Pure Science, University of Basra in the month of May, 2012 and the plant was botanically authenticated and voucher specimens (4124) were deposited in the Herbarium of Basra (Iraq, Basra, College of Science, University of Basra). The method of Lohachoompol et al. [
Extraction of anthocyanin from I. herbstii flower (AEIH) was carried out by using the method of [
The total anthocyanin content was measured using a modified pH differential method described by Lee et al. [
where A = [(A510 − A700) pH 1.0 ? (A510 − A700) pH 4.5]; Mw (molecular weight) = 449.2 g/mol for cyanidin-3- glucoside (cyd-3-glu); DF = dilution factor; ɛ = the molar absorptivity (26,900) and l = path length in cm. This assay was done in triplicate and average values were taken.
Total antioxidant activity of I. herbstii flower anthocyanin extract (AEIH) was determined by the method of Orak [
where Aο was the absorbance of control (blank, without extract) and A was the absorbance in the presence of the extract. This assay was done in triplicate and average values were taken.
The reducing power of I. herbstii flower anthocyanin extract (AEIH) was determined according to the method of Oyaizu [
Superoxide radicals of I. herbstii flower anthocyanin extract (AEIH) were generated by the method of Siddhurajuna et al. [
The hydroxyl radical scavenging activity of I. herbstii flower anthocyanin extract (AEIH) was determined according to the methods described by Singh et al. [
Hydrogen peroxide scavenging activity of I. herbstii flower anthocyanin extract (AEIH) was determined using the method described by Akinpelu et al. [
Nitric oxide generated from sodium nitroprosside was measured by the Griess Illosvoy reagent method Susuanta et al. [
The chelating of ferrous ions by the I. herbstii flower anthocyanin extract (AEIH) was estimated by the method of Ponmozhi et al. [
Inhibitory effect of I. herbstii flower anthocyanin extract (AEIH) on deoxyribose degradation was determined and hydroxyl radicals (referred to as non-site-specific scavenging assay) or antioxidants and iron ions (referred to as site-specific scavenging assay), described by Lee et al. [
The data were expressed as mean values ± SEM and tested with analysis of variance followed by Dunnett’s t-test. P-values < 0.05, 0.01 were considered to be statistically significant.
The total anthocyanins contents in fresh flowers after drying and during the 3 months storage are shown in
Treatments | Symbol | Total anthocyanin mg/g dry matter | % loss of anthocyanin |
---|---|---|---|
Fresh flower | AEFF | 8.31 ± 0.23 | ― |
Dried flower | AEDF | 7.17 ± 0.12* | 13.72 |
Flower 2-week storage at 5˚C | AESF | 6.43 ± 0.24** | 22.63 |
Flower stored frozen for 1 month | AEZF1 | 5.65 ± 0.33** | 32.01 |
Flower stored frozen for 3 month | AEZF3 | 4.71 ± 0.51** | 43.33 |
Total anthocyanin as cyanidin-3-glucoside equivalent, values are mean ± SEM, *P < 0.05, **P < 0.01, Dunnet test as compared to fresh flower group.
degradation was proportional depending on the percentage of soft flowers in the whole sample. Anthocyanins contents of frozen samples for 1 month (AEZF1) and 3 month (AEZF3) were found significantly decreased (P < 0.01) with compared to fresh sample (AEFF). This indicates that anthocyanins degradation rate was accelerated at longer storage. Probably, significant decrease of investigated compounds was due to water content in non- frozen state. Activity and enzymatic reaction rate reached maximum values in the layers of liquid water in frozen fruits. Perhaps, this phenomenon contributes to the modification of chemical compounds, including biologically active substances. In frozen products the enzymatic reactions are slow, but not completely blocked [
The total antioxidant activity of I. herbstii flower anthocyanin extract (AEIH) was determined by peroxidation of linoleic acid using ferric thiocyanate method. During linoleic acid peroxidation, peroxides were formed and these compounds oxidized Fe2⁺ to Fe3⁺. The Fe3⁺ ion forms a complex with SCN⁻, which had a maximum absorbance at 500 nm [
Furthermore, the reducing power of the extractable anthocyanins from I. herbstii flower (AEIH) is presented in
extracts to reduce Fe3+ to Fe2+ is a potent antioxidation defense mechanism, and two mechanisms available to effect this reducing power is by electron transfer and hydrogen atom transfer [
Superoxide anion radicals are produced by a number of cellular reactions, including various enzymes systems such as lipooxygenase, peroxidase, NADPH oxidase and xanthain oxidase. Superoxide anions place an important role in plant tissue and are involved in the formation of other cell damaging free radicals [
The hydroxyl radical is extremely reactive free radicals formed in biological system and has been implicated as a higher damaging species in free radical pathology. Capable of damaging almost every molecule found in living cells. This species is considered to be one of the quick initiators of the lipid peroxidation process, abstracting hydrogen atoms from unsaturated fatty acids [
process of lipid peroxidation. The anthocyanin extract (AEFF) seems to be a good scavenger of active oxygen species, thus reducing rate of chain reaction.
The measurement of H2O2-scavenging activity is one of the useful methods of determining the ability of antioxidants to decrease the level of pro-oxidants such as H2O2 [
gen peroxide could be an efficient assessment method to evaluate antioxidant property of I. herbstii flower anthocyanin extract (AEIH).
Nitric oxide [NO] radical is generated from sodium nitroprusside at physiological pH. It is a highly reactive compound that is capable of changing the structural and functional behavior of many cellular components [
Minimizing Fe2+ may afford protection against oxidative damage by inhibiting the production of ROS and lipid peroxidation [
Hydroxyl radicals can be formed by the Fenton’s reaction in the presence of reduced transition metal such as Fe2+ and H2O2, which is known to be the most reactive of all reduced from of dioxygen and thought to initiate all
damage in vivo [
From the results obtained in the present study, it may be concluded that the flowers of Iresine herbstii L. could
be evaluated as a majour source of anthocyanin. The amount of total anthocyanin was dramatically losses during vacuum drying, storage and freezing with compared to the fresh samples. The antioxidant profile of these compounds can be harnessed to treat radical related pathological conditions. The mechanism of antioxidant action was based on the ability of its extracts to donate electrons, reduce ferric ions, and scavenge superoxide anion, nitric oxide, hydrogen peroxide, and hydroxyl radicals. Thus this study gives support for expanding future investigations of pharmacological activities associated with free radicals and characterization of potent extract for its main active constituents.