Anna Rzepecka-Stojko

Polyphenols from Bee Pollen: Structure, Absorption, Metabolism and Biological Activity

Bee pollen constitutes a natural source of antioxidants such as phenolic acids and flavonoids, which are responsible for its biological activity. Research has indicated the correlation between dietary polyphenols and cardioprotective, hepatoprotective, anti-inflammatory, antibacterial, anticancerogenic, immunostimulating, antianaemic effects, as well as their beneficial influence on osseous tissue. The beneficial effects of bee pollen on health result from the presence of phenolic acids and flavonoids which possess anti-inflammatory properties, phytosterol and linolenic acid which play an anticancerogenic role, and polysaccharides which stimulate immunological activity. Polyphenols are absorbed in the alimentary tract, metabolised by CYP450 enzymes, and excreted with urine and faeces. Flavonoids and phenolic acids are characterised by high antioxidative potential, which is closely related to their chemical structure. The high antioxidant potential of phenolic acids is due to the presence and location of hydroxyl groups, a carboxyl group in the immediate vicinity of ortho-diphenolic substituents, and the ethylene group between the phenyl ring and the carboxyl group. As regards flavonoids, essential structural elements are hydroxyl groups at the C5 and C7 positions in the A ring, and at the C3′ and C4′ positions in the B ring, and a hydroxyl group at the C3 position in the C ring. Furthermore, both, the double bond between C2 and C3, and a ketone group at the C4 position in the C ring enhance the antioxidative potential of these compounds. Polyphenols have an ideal chemical structure for scavenging free radicals and for creating chelates with metal ions, which makes them effective antioxidants in vivo.

Caffeic Acid Phenethyl Ester and Ethanol Extract of Propolis Induce the Complementary Cytotoxic Effect on Triple-Negative Breast Cancer Cell Lines

Chemotherapy of breast cancer could be improved by bioactive natural substances, which may potentially sensitize the carcinoma cells’ susceptibility to drugs. Numerous phytochemicals, including propolis, have been reported to interfere with the viability of carcinoma cells. We evaluated the in vitro cytotoxic activity of ethanol extract of propolis (EEP) and its derivative caffeic acid phenethyl ester (CAPE) towards two triple-negative breast cancer (TNBC) cell lines, MDA-MB-231 and Hs578T, by implementation of the MTT and lactate dehydrogenase (LDH) assays. The morphological changes of breast carcinoma cells were observed following exposure to EEP and CAPE. The IC50 of EEP was 48.35 µg∙mL−1 for MDA-MB-23 cells and 33.68 µg∙mL−1 for Hs578T cells, whereas the CAPE IC50 was 14.08 µM and 8.01 µM for the MDA-MB-231 and Hs578T cell line, respectively. Here, we report that propolis and CAPE inhibited the growth of the MDA-MB-231 and Hs578T lines in a dose-dependent and exposure time-dependent manner. EEP showed less cytotoxic activity against both types of TNBC cells. EEP and, particularly, CAPE may markedly affect the viability of breast cancer cells, suggesting the potential role of bioactive compounds in chemoprevention/chemotherapy by potentiating the action of standard anti-cancer drugs.

Antioxidative properties of bee pollen extracts examined by EPR spectroscopy

Antioxidative Properties of Bee Pollen Extracts Examined by EPR Spectroscopy Bee pollen is a valuable and highly recognized source of exogenous antioxidants. The aim of these studies was to determine the antioxidant capacity of three types of bee pollen extracts: ethanol extracts of bee pollen, pepsin extracts of bee pollen and ethanol extracts of pepsin-digested bee pollen. Their antioxidant properties were determined with the use of electron paramagnetic resonance (EPR) and their ability to quench DPPH free radicals was estimated. The EPR results showed that ethanol extracts of pepsin-digested bee pollen (EEPP) had the highest antioxidative effect and the highest free radical DPPH scavenging potential. The pepsin extracts of bee pollen (PEP) had the weakest antioxidant capacity. The ability to quench DPPH free radicals was also the weakest one for this extract. An average antioxidative effect was recorded for ethanol extracts of bee pollen (EEP). Właściwości Antyoksydacyjne Ekstraktów z Pyłku Pszczelego Badane Metodą Spektroskopii EPR W ostatnim czasie dużo uwagi poświęca się substancjom o działaniu antyoksydacyjnym pochodzenia naturalnego. Cennym i uznanym źródłem egzogennych antyoksydantów jest pszczeli pyłek kwiatowy. W związku z powyższym, przedmiotem badań było oznaczenie aktywności antyoksydacyjnej trzech rodzajów ekstraktów z pyłku pszczelego: ekstraktu etanolowego pyłku pszczelego, ekstraktu pepsynowego pyłku pszczelego oraz ekstraktu etanolowego po uprzednim trawieniu enzymatycznym pepsyną pyłku pszczelego. Właściwości antyoksydacyjne oznaczono metodą elektronowego rezonansu paramagnetycznego (EPR) na podstawie zdolności redukcji wolnego rodnika DPPH. Najwyższą aktywnością antyoksydacyjną oznaczoną metodą EPR charakteryzował się ekstrakt etanolowy po uprzednim trawieniu enzymatycznym pepsyną pyłku pszczelego (EEPP). Dla tego rodzaju ekstraktu zdolność redukcji wolnego rodnika DPPH była najsilniejsza. Z kolei ekstrakt pepsynowy pyłku pszczelego (PEP) cechował się najniższą aktywnością antyoksydacyjną. Zdolność redukcji wolnego rodnika DPPH dla tego ekstraktu była najsłabsza. Pośrednią aktywność antyoksydacyjną spośród badanych ekstraktów wykazano dla ekstraktu etanolowego pyłku pszczelego (EEP).

Biological Activity of Propolis-Honey Balm in the Treatment of Experimentally-Evoked Burn Wounds

Medicines of biogenic origin with micro-organic, regenerative and analgesic properties are becoming more and more significant in the treatment of burn wounds. These properties are found in apitherapeutics such as propolis and honey—products collected and processed by a honey bee. Their effect on the course of the healing processes is multidirectional. The aim of the study was a histopathological and biochemical analysis of the processes of scar formation in experimentally evoked burn wounds in white pigs treated with the 1% and 3% Sepropol balms containing standardized extracts of propolis and honey. The results were compared with the therapeutic effects obtained with dermazin cream (1% silver sulfadiazine). The level of collagen was determined in the wounds treated with 1% and 3% Sepropol and compared with the collagen level in healthy skin and wounds treated with dermazin. Granulation and regenerated epithelium formation times were compared, with the 3% Sepropol being by far the most effective. The 3% Sepropol also increased the collagen level to 116% with the control sub-groups scoring between 80% and 98%. The results show the healing process of burn wounds in pigs treated with the Sepropol balm starts earlier and has a faster course than the standard dermazin therapy.

Migration Rate Inhibition of Breast Cancer Cells Treated by Caffeic Acid and Caffeic Acid Phenethyl Ester: An In Vitro Comparison Study

One of the deadliest cancers among women is a breast cancer. Research has shown that two natural substances occurring in propolis, caffeic acid (CA) and caffeic acid phenethyl ester (CAPE), have significant anticancer effects. The purpose of our in vitro study was to compare cytotoxic activity and migration rate inhibition using CA and CAPE (doses of 50 and 100 µm) against triple-negative, MDA-MB-231 breast adenocarcinoma line cells, drawn from Caucasian women. Viability was measured by XTT-NR-SRB assay (Tetrazolium hydroxide-Neutral Red-Sulforhodamine B) for 24 h and 48 h periods. Cell migration for wound healing assay was taken for 0 h, 8 h, 16 h, and 24 h periods. CAPE displayed more than two times higher cytotoxicity against MDA-MB-231 cells. IC50 values for the XTT assay were as follows: CA for 24 h and 48 h were 150.94 µM and 108.42 µM, respectively, while CAPE was 68.82 µM for 24 h and 55.79 µM for 48 h. For the NR assay: CA was 135.85 µM at 24 h and 103.23 µM at 48 h, while CAPE was 64.04 µM at 24 h and 53.25 µM at 48 h. For the SRB assay: CA at 24 h was 139.80 µM and at 48 h 103.98 µM, while CAPE was 66.86 µM at 24 h and 47.73 µM at 48 h. Both agents suspended the migration rate; however, CAPE displayed better activity. Notably, for the 100 µM CAPE dose, motility of the tested breast carcinoma cells was halted.

Comparison of Two Components of Propolis: Caffeic Acid (CA) and Caffeic Acid Phenethyl Ester (CAPE) Induce Apoptosis and Cell Cycle Arrest of Breast Cancer Cells MDA-MB-231

Studies show that caffeic acid (CA) and caffeic acid phenethyl ester (CAPE) are compounds with potent chemopreventive effects. Breast cancer is a common form of aggressive cancer among women worldwide. This study shows a comparison of CA and CAPE activity on triple-negative human caucasian breast adenocarcinoma line cells (MDA-MB-231). MDA-MB-231 cells were treated by CA and CAPE with doses of from 10 to 100 µM, for periods of 24 h and 48 h. Cytotoxicity MTT tests, apoptosis by Annexin V, and cell cycle with Dead Cell Assays were performed. Cytotoxic activity was greater for CAPE compared to CA (both incubation times, same dosage). IC50 values for CAPE were 27.84 µM (24 h) and 15.83 µM (48 h) and for CA > 10,000 µM (24 h) and > 1000 µM (48 h). Polyphenols induced apoptosis, while CAPE (dose dependently), induced a higher apoptotic effect. CAPE also induced cell cycle arrest in S phase (time and dose dependently), CA did it only for 50 and 100 µM. A dose dependent decline was seen for the G0/G1 phase (CAPE, 48 h), as well as elimination of phase G2/M by 100 µM of CAPE (only mild effect for CA). Comparing CA and CAPE activity on MDA-MB-231, CAPE clearly showed better activity for the same dosages and experiment times.

Antioxidant activity of ethanolic fractions of Polish propolis.

There is a great variation in the chemical composition of propolis of different origins. Likewise, the method of its extraction has signifi cant impact on the content of biologically active compounds. Here we compared methods of propolis extraction for optimal antioxidant activities which were measured by means of β-carotene discolouration, 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging, and 2,2’-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS•+) radical cation decolouration assays. In the extracts, the contents of polyphenols and fl avonoids were measured, and phenolic acids were identifi ed and quantifi ed by HPLC. A three-step extraction allowed obtaining large amounts of phenolic acids from propolis. The propolis fractions obtained had antioxidant properties comparable to those of α-tocopherol and butylated hydroxytoluene. Therefore, they may be used as effective natural antioxidants.

Polyphenol content and antioxidant activity of bee pollen extracts from Poland

Our study determined the effect of extraction method of bee pollen on the optimal antioxidant activity of the extract obtained. We determined the content of polyphenols and flavonoids as well as the antioxidant activity of the ethanol extract of pollen loads (EEP), the pepsin extract of pollen loads (PEP), and the pepsin-digested extract of pollen loads (EEPP). The total content of polyphenols was measured using Follin–Ciocalteau reagent. The flavonoid content was measured using aluminum chloride. Phenolic acids and flavonoids were identified and quantified by HPLC. The antioxidant activity was measured by 2,2-diphenyl-picrylhydrazyl radical scavenging activity assays and Trolox equivalents antioxidant capacity. Antioxidant activities were the highest in EEPP and associated with the total content of phenolic and flavonoid compounds. This study indicated that pepsin digestion conducted before ethanol extraction allowed us to obtain more bioactive compounds, as well as the highest antioxidant activity of extract.

Anti-Atherogenic Activity of Polyphenol-Rich Extract from Bee Pollen

The aim of this study was to determine the effect of polyphenol-rich ethanol extract of bee pollen (EEP) on atherosclerosis induced by a high-fat diet in ApoE-knockout mice. EEP was given with feed in two doses of 0.1 and 1 g/kg body mass (BM). The studies have been conducted in a period of 16 weeks. The following factors were estimated: total cholesterol (TC), oxidized low density lipoproteins (ox-LDL), asymmetric dimethylarginine (ADMA), angiotensin-converting enzyme (ACE) and angiotensin II (ANG II) in the 5th, 10th, 12th, 14th, and 16th week of the experiment. In the last, i.e., 16th week of the studies the development of coronary artery disease (CAD) was also estimated histopathologically. Supplementing diet with EEP resulted in decreasing TC level. EEP reduced oxidative stress by lowering the levels of ox-LDL, ADMA, ANG II and ACE. EEP protected coronary arteries by significantly limiting the development of atherosclerosis (the dose of 0.1 g/kg BM) or completely preventing its occurrence (the dose of 1 g/kg BM). The obtained results demonstrate that EEP may be useful as a potential anti-atherogenic agent.

Protective effects of vitamin E against CCl4-induced hepatotoxicity in rabbits

The influence of CCl4 on the activity of superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR), the value of the total antioxidant status (TAS), and the concentration of malonic dialdehyde (MDA) and glutathione (GSH) was monitored in plasma or whole blood of rabbits. The administration of CCl4 caused the increase of the SOD activity to approximately 150 % and the decrease in the activity of GPx and GR by about 50 %. These changes were accompanied with the increase in TAS value and MDA concentration and the decrease of GSH concentration. The effect of CCl4 was suppressed by the previous 7 days lasting or simultaneous administration of vitamin E. Oxidative stress caused by CCl4 was accompanied by the development of reactive oxygen forms, especially superoxide radical anion.