Cancer is a disease caused by uncontrolled cell growth. Cancer treatments, such as chemotherapy, have tended to cause adverse effects for patients. An alternative that can be described is the development of natural-based cancer therapies that have relatively low side effects. The development of Na-alginate as a matrix for nanoencapsulation of Uncaria gambir has shown growth inhibition activity against HeLa and T47D cell lines. The EC50 ± SE of U. gambir extract on the HeLa cell line was 1055.14 ± 42.52 µg/mL while on the T47D cell line 297.15 ± 15.41 µg/mL. On the other hand, the Ug-NPs anticancer activity was substantially higher with EC50 ± SE for HeLa cell line 28.12 ± 27.10 µg/mL, and T47D cell line 10.39 ± 3.50 µg/mL. Therefore, the nanoencapsulation of U. gambir extracts has increased solubility, selectivity, and activity.
Article
Anticancer Activity of Nano Uncaria gambir
https://doi.org/10.21203/rs.3.rs-1497841/v1
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Anticancer
HeLa cells
nanoencapsulation
Na-alginate
T47D cells
Uncaria gambir
Cancer is an uncontrolled and abnormal proliferation of various cells in the body. Moreover, there are 100 different cancers, which vary substantially in action and response to treatment1. The most common cancer diagnosed in both men and women is lung cancer (11.6% of the total cases). The leading cause of cancer death (18.4% of total cancer deaths). This is followed by breast (11.6%), colorectal (9.2%), gastric (8.2%), and liver cancers (8.2%) 2. In Southeast Asia, Indonesia (136.2 / 100,000 population) ranks 8th in cancer prevalence, while in Asia, it is 23rd. Based on RISKESDAS data, the prevalence of tumors or cancer in Indonesia increased from 1.4 per 1000 population in 2013 to 1.79 per 1000 population in 2018 3.
Treatment of cancer so far is based on radiotherapy and chemotherapy. The adverse effects of both are often devastating because, in killing cancer cells, these two methods can also harm normal cells 4. The adverse chemotherapy effects include hair loss, bone marrow suppression, drug resistance, gastrointestinal lesions, neurological dysfunctions, and cardiac toxicity 5. Besides, Herbal-based medicines are perceived to have fewer side effects than chemical drugs. It has triggered the development of herbal-based drugs increasingly becoming the focus for treatment regimens 6. The cancer treatment with herbs can inhibit or reduce cancer cell proliferation and help control the adverse effect of conventional 7,8.
U. gambir is a species belonging to the Rubiaceae family. This species is found in Sumatra as a leading commodity from the West Sumatra province 8. Catechins are among the bioactive components of U. gambir 9. Hence, Catechins have better T47D cell line inhibitory activity than MCF7 cell lines 10. Anticancer activity of catechins is based on suppressing cell proliferation by inducing apoptosis, inhibiting angiogenesis, and metastasis of cancer cells 11. Hydrated catechins exhibit anticancer effects by blocking MCF7 cell proliferation and inducing apoptosis by modulating the expression levels of caspase-3, -8, -9, and p53 12.
The nanotechnology approach was applied for cancer chemotherapy to reduce toxicity, and increase stability, bioavailability, and selectivity 13. Nanoencapsulation is one of the nanotechnological applications aimed to increase the stability and bioavailability of bioactive compounds because nano-size can increase the surface area of an active ingredient 14,15. Nanoscale drug delivery systems are formulated from biocompatible and biodegradable polymers. They are being used to develop the administration and targeting strategy for cancer drugs 16. Nanoencapsulation of U. gambir using sodium alginate to increase the plant's anticancer activity.
Material
Sodium alginate food grade (Na-Alginate), distilled water, methanol, ethyl acetate, n-hexane, phosphate buffer, ethanol absolute (Merck), phosphate buffer saline (PBS); HCl (Sigma-Aldrich); NaCl (Sigma-Aldrich); NaOH (Sigma-Aldrich); sodium dodecyl sulfate (SDS), and 3-(4,5'-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT). The U. gambir sap was obtained from traditional markets in Surabaya, Indonesia.
Extraction of U. gambir
The dried U. gambir sap (500 g) was extracted by the maceration method using methanol as a solvent with a ratio of 1: 2 at room temperature for 24 hours. The methanol extract was then concentrated using a rotary vacuum evaporator. This extract was then partitioned with n-hexane and ethyl acetate. Ethyl acetate extract was concentrated using a rotary vacuum evaporator.
Determination of Catechin from U. gambir
Methanol, ethyl acetate, and hexane extracts of U. gambir were analyzed by UV-Vis spectrometer for catechin content. The extract with the highest catechin content was selected for nanoencapsulation.
Synthesis of Ug-NPs 17
The U. gambir ethyl acetate extract was added with Na-alginate solution in a ratio of 1 : 2. The solution mixture was homogenized using ultrasonic (JY-9211DN, Ningbo Scientz Biotechnology, China) to obtain Ug-NPs. The Ug-NPs functional group was characterized by FTIR (Shimadzu IRTracer-100). The physicochemical properties of Ug-NPs, including polydispersity index (PDI), particle size, zeta potential (ζ) (Dynamic Light Scattering, Zetasizer Nano ZS, Malvern). Atomic Force Microscope (AFM, Agilent Technologies 5500) was determined for sample morphology, and TGA (Perkin Elmer TGA 4000) was used to observe the decomposition changes of Ug-NPs.
The Analysis of Ug-NPs Stability 17
The stability test was aimed to determine the Ug-NPs stability, which is influenced by pH, temperature, and NaCl concentration. The observation was made by changing UV-Vis spectra and the level of turbidity of Ug-NPs.
Loading and Release of Ug-NPs 17,18
The Calculations of Loading Efficiency (LE) and Loading Amount (LA) were based on equations (1) and (2). The loading value interpreted how much of the bioactive components of U. gambir extract were trapped in the Ug-NPs.
The release of material was determined as the amount of catechins from U. gambir extract that could be released at pH 4, 7, and 9.
Cytotoxicity Test
HeLa and T47D cell lines that have been grown on RPMI 1640 media and have 80% confluence conditions are distributed on 96-well plates with several 10 x 104 cells / well. Cells were incubated for 24 hours in a CO2 incubator, then 100 µL of culture media containing samples (U. gambir extract and Ug-NPs) were added and incubated again for 24 hours. At the end of incubation, the culture medium in the plate was discarded and then washed with PBS. Each well was added with 100 µL of MTT (5 mg/mL) reagent, incubated again for 4 hours at 37°C in 5% CO2. After 4 hours, 100 µL of 10% SDS solution was added in 0.01 N HCl. The absorbance was measured using an ELISA reader (Bio-Rad) at λ 550 nm. The EC50 values were obtained by using a dose-response calculation.
Statistical Analysis
One-way ANOVA analysis was used to determine the anticancer effect of U. gambir extract with Ug-NPs on HeLa and T47D cell lines. Post-hoc analysis was carried out by Tukey’s test. If the p-value was < 0.05, then statistically U. gambir nanoencapsulation process significantly increased the anticancer activity.
Catechins of U. gambir
U. gambir sap was extracted with methanol and partitioned with n-hexane and ethyl acetate. Catechin levels in the three U. gambir extracts are shown in Fig. 1. U. gambir ethyl acetate extract has the highest catechin content, 89.34% (w/w), then processed for nanoencapsulation with Na-alginate. Catechins are bioactive compounds that have been reported for anticancer activity.
Ug-NPs Characterization
Ug-NPs is a nano-capsule of U. gambir extract with Na-alginate. Nano-capsule characterization based on physicochemical properties is shown in Table 1. It was injured that Ug-NPs have a particle size of 452.63 ± 5.29 nm based on the particle size. The polydispersity index (PDI) measures size heterogeneity that indicates size distribution or agglomeration/aggregation in sample 19. PDI Ug-NPs value 0.55 ± 0.01 indicates that the particle size distribution tends to be homogeneous. The particle size distribution is considered broad if the PDI > 0.7 20.
| Type | Size ± SD (nm) | PDI ± SD | ζ ± SD (mV) |
|---|---|---|---|
| Na-Alginate | 600.20 ± 105.02 | 1.00 ± 0.00 | -37.20 ± 3.06 |
| U. gambir extract | 586.00 ± 13.86 | 0.47 ± 0.05 | -44.47 ± 0.47 |
| Ug-NPs | 452.63 ± 5.29 | 0.55 ± 0.01 | -40.87 ± 0.90 |
| Notes: Each data presented as mean ± SD (n = 3) | |||
The smaller the polydispersity index number, the more uniform the particle size because if the size difference between particles is bigger, this will affect the particle characteristics 21. Zeta potential (ζ) in Table 1 shows that Na-alginate, U. gambir extract, and Ug-NPs are negative. This may be due to the large number of hydroxy groups that are electronegative. The Zeta potential value for Ug-NPs is between Na-alginate and U. gambir extract. This is presumably because Na-alginate's electropositive group partially covers the electronegative group in the constituent U. gambir extract. Nanoencapsulation with a zeta potential value of more than ± 30 mV is stable in suspension as surface charge prevents aggregation 22. A large Zeta potential indicates better stability due to the larger electrostatic repulsion between particles. The Ug-NPs tend to be stable with a -40.87 ± 0.90 Mv value. Morphological analysis by AFM showed that the Ug-NPs were spherical (Fig. 2).
FTIR spectra of Ug-NPs showed a blend of Na-alginate and U. gambir extract peaks. Absorptions at 1519 cm− 1 (C-C in ring) and 1033 cm− 1 (C-O-C stretching vibration) were from U. gambir extract. Meanwhile, absorption at 1419 cm− 1 (vibrations of carboxylate salt ion) and 1122 cm− 1 (deformation -OH in -COOH) were from Na-alginate (Fig. 3).
The TGA analysis of Ug-NPs was carried out to determine the composition and the interaction between the active compounds in the U. gambir extract and the sodium alginate coating material (Fig. 4). The Ug-NPs decomposed at a temperature of 70o C, which showed a decrease in sample weight of ± 10.5%. The U. gambir extract also decomposed at a temperature of around 70o C with a decrease in sample weight of ± 12.1%. At a temperature of 178o C, Ug-NPs and extracts of U. gambir decomposed with a decrease in sample weight of ± 16.2% and ± 33.1%, respectively. The Na-alginate decomposed at a temperature of 290o C with a decrease in sample weight of ± 35.3%. At the same temperature, Ug-NPs also decomposed with a decrease in sample weight of ± 25.2%. Based on FTIR and TGA analysis, it is suspected that only physical interactions exist between the constituent of U. gambir extract and Na-alginate in the Ug-NPs, and apparently, no chemical interaction exists between the two.
Stability of Ug-NPs
The stability of Ug-NPs must be maintained so that there is no aggregation and changes in the bioactive components of Ug-NPs. The UV-Vis spectra on the Ug-NPs showed two absorption bands indicating bioactive catechins in U. gambir extract. Change in pH from 3 to 11 tended had to affect no the absorption bands of the Ug-NPs, which ranged from 383 to 386 nm. At pH 12, there was a bathochromic shift in the first absorption band to 410 nm. In general, in the second absorption band, changes in pH from 3 to 11 result in a hyperchromic effect. Meanwhile, at pH 12, it causes a hypsochromic and hyperchromic shift. The turbidity level of Ug-NPs tends to be higher at acidic pH 3–5 and lower at pH 6–12. It is influenced by the stability of Na-alginate, which tends to be less stable at acidic pH. Consequently, the Na-alginate precipitates. Below pH 5, the free –COO- ions will form protonated -COOH. As a result, the electrostatic repulsion between the chains decreases, and they can approach and form hydrogen bonds. This increases in viscosity. Meanwhile, if the pH is more alkaline, the depolymerization is slow. Consequently, the viscosity will decrease 23,24 (Supplementary Fig. 1).
The temperature changes between 30o C − 100o C generally did not affect the UV-Vis absorption pattern and did not significantly affect the turbidity level of Ug-NPs. Increasing the temperature will decrease the viscosity of Na-alginate (Supplementary Fig. 2) 24. Ionic stability of Ug-NPs showed that NaCl concentration did not significantly affect the UV-Vis absorption band of Ug-NPs, both at 0–0.3 M concentrations. However, increasing NaCl concentration affected the turbidity of Ug-NPs. The higher concentration of NaCl is associated with the higher turbidity levels of Ug-NPs. It may be due to the stability of Na-alginate in NaCl solution. Monovalent salt (NaCl) can affect the rate of hydration of Na-alginate, which results in precipitation (Supplementary Fig. 3) 25.
Loading and Release of Ug-NPs
Loading efficiency (LE) showed that 97.56 ± 0.04% of the bioactive components of U. gambir extract are adsorbed in Na-alginate micelles. Meanwhile, 32.52 ± 0.01% is the loading amount (LA) of the bioactive components of U. gambir extract in Ug-NPs. The release of bioactive components of U. gambir extract on the Ug-NPs showed that at pH 4 the release tends to be slow. It showed that the Ug-NPs stability at pH 4, is less stable due to a decrease in the electrostatic repulsion of Na-alginate. Thus, it will result in the Ug-NPs aggregation. Besides, the formation of hydrogen bonds results in the shrinking of the pores of the Ug-NPs. So, the release tends to be slower i.e. 12.10 ± 0.03% after 24 hours. For pH 7 and 9, the release tends to be faster i.e. 27.65 ± 0.03% and 31.37 ± 0.00% after 24 hours, respectively. This is because the repulsion between particles at neutral or alkaline pH causes the pores of the Ug-NPs to enlarge, which results in a faster release of bioactive components (Fig. 5).
Cytotoxic Activity of Ug-NPs
The anticancer activity of U. gambir extract, and Ug-NPs was evaluated against HeLa and T47D cell lines and are presented as EC50 ± SE. In the HeLa cell lines, U. gambir extract was less active in inhibiting the cell growth with EC50 1055.14 ± 42.52 µg/mL. Meanwhile, the Ug-NPs significant EC50 28.12 ± 27.10 µg/mL (Fig. 6A). It was supported by statistical analysis, which showed a significant p = 0.000. It indicated a substantial increase in the activity of U. gambir extracts after nanoencapsulation.
On the T47D cell line, the U. gambir extract showed EC50 297.15 ± 15.41 µg/mL, while Ug-NPs was EC50 10.39 ± 3.50 µg/mL (Fig. 6B). It indicates that the U. gambir extract nanoencapsulation has increased the growth inhibitory activity of the T47D cell line. The statistical test showed that U. gambir extract nanoencapsulation significantly increased the growth inhibitory activity with a p-value = 0.003.
U. gambir is among the plants with high catechin content. The catechin content of U. gambir ethyl acetate extract was 89.34%. One of the bioactivities of catechins is as an anticancer. The mechanism of anticancer activity of catechins can be through diverse mechanisms such as antioxidants 26, regulation of drug-metabolizing enzymes, induction of apoptosis 27, inhibition of cell proliferation 28, anti-inflammatory 29, inhibiting cancer metastasis 30, and regulation of gut microbiota 31,32. Even though they have high bioactivity, natural ingredients tend to have low solubility and are non-specifically. Such factors affect their overall bioactivity 33.
U. gambir nanoencapsulation process can enhance its anticancer bioactivity. Nanoencapsulation can increase the bioavailability of active plant compounds. The absorption surface area is large in smaller particle sizes and can effectively participate in a paracellular pathway and enter the systemic channel 34. This process can maximize the bioactivity of U. gambir extract. The advantages of Na-alginate as a coating material for U. gambir nano-capsules are biodegradable, non-toxic, biocompatible, and non-mutagenic 35. Na-alginate also plays an essential role in drug delivery of chemotherapy drug components, so it is appropriate for use in the nanoencapsulation of U. gambir as chemotherapy 36,37.
Nanoencapsulation of U. gambir extract using Na-alginate was proven to prevent damage or degradation of bioactive components. It was inferred from the UV-Vis spectra of the Ug-NPs, which tend to be stable at various pH, temperature, and salt concentrations. Additionally, the Ug-NPs can control the release of bioactive components, increase the dissolution rate and permeability of bioactive components of U. gambir, prolong plasma half-life, and improve pharmacokinetic profile 38.
Ug-NPs proved to be successful in increasing the anticancer activity of U. gambir. The growth inhibitory activity against HeLa cells increased by 97.33%, while in the case of T47D cells, it is 96.50%. The increase in the anticancer activity of the Ug-NPs against the HeLa and T47D cell lines was due to the ability of the Ug-NPs to deliver bioactive components of U. gambir across the plasma membrane. The plasma membrane is an effective barrier in protecting all living cells from the surrounding environment. Additionally, it firmly controls the entry and exit of macromolecular substances. Therefore, a nano-capsule system is needed to overcome this barrier to enter into living cells 39.
When reaching the cell exterior membrane, Ug-NPs can interact with the plasma membrane or the extracellular matrix components and enter the cell, mainly through the process of endocytosis 40. Cellular uptake of the Ug-NPs is measuring more than 200 nm. It is through one of the endocytosis pathways, called macro-pinocytosis 41. In the process of macro-pinocytosis, all particles and molecules dissolved in the extracellular fluid are captured by a membrane extension or ruffles. Then, it forms macro-pinosomes with a diameter of 0.2-5 µm [40]. After internalization of Ug-NPs into cells, macro-pinosomes fuse with lysosomes 42 and the Na-alginate matrix of the Ug-NPs is degraded by lysozyme, resulting in a release of bioactive components U. gambir extract into cells 43.
The charge on the surface determines the cellular internalization of the nano-capsule. Moreover, the positive charged nano-capsules are rapidly internalized into cells via the clathrin-dependent endocytosis pathway 41. The Ug-NPs with a positive charge can form clathrin vesicles. The vesicles separate from the plasma membrane and enter into the cytoplasm. Then, they diffuse into the endosome/lysosomal compartment. The environmental acidity increases the breakdown of bonds, and the chemical structures of the Ug-NPs by enzymes, causing rapid release 39. Additionally, positive charge nano-capsules induce membrane depolarization in several different cell types. Membrane depolarization produces an increased concentration of Ca2+ in the cell that can induce modulation of the intracellular pathway 40.
U. gambir contains catechin compounds that are known for anticancer activity. Nanoencapsulation of U. gambir extract can increase its anticancer activity. The Ug-NPs formulation has exhibited a better growth inhibitory activity than U. gambir extract against HeLa and T47D cell lines. The nanoencapsulation process of U. gambir extract with Na-alginate can also increase the drug delivery in cells, increasing their bioactivity.
Acknowledgments
Thanks to the L.E.J Nanotechnology Center, International Center for Chemical and Biological Science (ICCBS) for assisting in AFM based analysis.
Funding
This research was supported by the Directorate of Research and Community Service, Ministry of Research, Technology, and Higher Education through the "Penelitian Terapan Unggulan Perguruan Tinggi (PTUPT)" scheme in 2021 (Contract number: 456/UN3.15/PT/2021).
Availability of data and materials
Not applicable
Author’s contributions
APW, NSA, HIZ, MZF and SHL synthesized Ug-NPs. ALF, and AY tested the anticancer activity. MIC helped in the characterizations of Ug-NPs. The NSA wrote the manuscript, assisted by APW, SHL, and HIZ. ALF, AY, MZF, and MIC kept records and revise manuscripts. The NSA finalized the manuscript. All authors have approved the final manuscript.
Consent of publication
Not applicable
Competing interests
In this study, the author has no competing interest.
- Cooper GM, Hausman R. A molecular approach. The Cell. 2nd ed. Sunderland, MA: Sinauer Associates. 2000.
- Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA: Cancer J Clin. 2018;68(6):394–424.
- Kemenkes. Hari Kanker Sedunia 2019. 2019; http://www.depkes.go.id/article/view/19020100003/hari-kanker-sedunia-2019.html. Accessed July 05 2019
- Liu H, Lv L, Yang K. Chemotherapy targeting cancer stem cells. Am J Cancer Res. 2015;5(3):880.
- Hosseini A, Ghorbani A. Cancer therapy with phytochemicals: evidence from clinical studies. Avicenna J Phytomed. 2015;5(2):84–97.
- Greenwell M, Rahman P. Medicinal plants: their use in anticancer treatment. Int J Pharm Sci Res. 2015;6(10):4103–4112.
- Du G-J, Zhang Z, Wen X-D, et al. Epigallocatechin gallate (EGCG) is the most effective cancer chemopreventive polyphenol in green tea. Nutrients. 2012;4(11):1679–1691.
- Phillipson J, CE R. Alkaloids of Uncaria. V. their occurrence and chemotaxonomy. 1978.
- Anggraini T, Tai A, Yoshino T, Itani T. Antioxidative activity and catechin content of four kinds of Uncaria gambir extracts from West Sumatra, Indonesia. Afr J Biochem Res. 2011;5(1):33–38.
- Evacuasiany E, Ratnawati H, Liana LK, et al. Cytotoxic and antioxidant activities of catechins in inhibiting the malignancy of breast cancer. Oxid Antioxid Med Sci. 2014;3(2):141–146.
- Yu Y, Deng Y, Lu B-m, Liu Y-x, Li J, Bao J-k. Green tea catechins: a fresh flavor to anticancer therapy. Apoptosis. 2014;19(1):1–18.
- Alshatwi AA. Catechin hydrate suppresses MCF-7 proliferation through TP53/Caspase-mediated apoptosis. J Exp Clin Cancer Res. 2010;29(1):1–9.
- Bharali DJ, Mousa SA. Emerging nanomedicines for early cancer detection and improved treatment: current perspective and future promise. Pharmacol Ther. 2010;128(2):324–335.
- Shamsara O, Muhidinov ZK, Jafari SM, et al. Effect of ultrasonication, pH and heating on stability of apricot gum–lactoglobuline two layer nanoemulsions. Int J Biol Macromol. 2015;81:1019–1025.
- Mehrnia M-A, Jafari S-M, Makhmal-Zadeh BS, Maghsoudlou Y. Crocin loaded nano-emulsions: factors affecting emulsion properties in spontaneous emulsification. Int J Biol Macromol. 2016;84:261–267.
- Haley B, Frenkel E. Nanoparticles for drug delivery in cancer treatment. Urol Oncol. 2008;26(1):57–64.
- Wardana AP, Aminah NS, Fahmi MZ, et al. Nanoencapsulation of Syzygium polycephalum Extract Using Folate Modified κ-Carrageenan as Vehicles for Pronounced Anticancer Activity. Trop J Nat Prod Res. 2020;4(11):945–952.
- Fahmi MZ, Haris A, Permana AJ, et al. Bamboo leaf-based carbon dots for efficient tumor imaging and therapy. RSC Adv. 2018;8(67):38376–38383.
- Mudalige T, Qu H, Van Haute D, Ansar SM, Paredes A, Ingle T. Characterization of Nanomaterials: Tools and Challenges. Nanomaterials for Food Applications: Elsevier; 2019:313–353.
- Worldwide MI. Dynamic light scattering, Common terms defined. Inform white paper. Malwern Instruments Limited. 2011;2011:1–6.
- Manmode AS, Sakarkar DM, Mahajan NM. Nanoparticles-tremendous therapeutic potential: a review. Int J Pharmtech Res. 2009;1(4):1020–1027.
- Mohanraj V, Chen Y. Nanoparticles-a review. Trop J Pharm Res. 2006;5(1):561–573.
- Lee KY, Mooney DJ. Alginate: properties and biomedical applications. Prog Polym Sci. 2012;37(1):106–126.
- Qin Y. Seaweed hydrocolloids as thickening, gelling, and emulsifying agents in functional food products. Bioactive Seaweeds for Food Applications: Elsevier; 2018:135–152.
- McHugh DJ. Production, properties and uses of alginates. Production and Utilization of Products from Commercial Seaweeds. FAO. Fis. Tech Pap. 1987;288:58–115.
- Khan N, Afaq F, Saleem M, Ahmad N, Mukhtar H. Targeting multiple signaling pathways by green tea polyphenol (–)-epigallocatechin-3-gallate. Cancer Res. 2006;66(5):2500–2505.
- Pan M-H, Chiou Y-S, Wang Y-J, Ho C-T, Lin J-K. Multistage carcinogenesis process as molecular targets in cancer chemoprevention by epicatechin-3-gallate. Food Funct. 2011;2(2):101–110.
- Crozier A, Jaganath IB, Clifford MN. Dietary phenolics: chemistry, bioavailability and effects on health. Nat Prod Rep. 2009;26(8):1001–1043.
- Mackenzie GG, Carrasquedo F, Delfino JM, Keen CL, Fraga CG, Oteiza PI. Epicatechin, catechin, and dimeric procyanidins inhibit PMA-induced NF‐κB activation at multiple steps in Jurkat T cells. FASEB J. 2004;18(1):167–169.
- Ko H, So Y, Jeon H, et al. TGF-β1-induced epithelial–mesenchymal transition and acetylation of Smad2 and Smad3 are negatively regulated by EGCG in human A549 lung cancer cells. Cancer Lett. 2013;335(1):205–213.
- Chen H, Sang S. Biotransformation of tea polyphenols by gut microbiota. J Funct Foods. 2014;7:26–42.
- Jiang Y, Jiang Z, Ma L, Huang Q. Advances in nanodelivery of green tea catechins to enhance the anticancer activity. Molecules. 2021;26(11):3301.
- Yao H, Liu J, Xu S, Zhu Z, Xu J. The structural modification of natural products for novel drug discovery. Expert Opin Drug Discov. 2017;12(2):121–140.
- Delie F, Blanco-Príeto MJ. Polymeric particulates to improve oral bioavailability of peptide drugs. Molecules. 2005;10(1):65–80.
- Chaturvedi K, Ganguly K, More UA, et al. Sodium alginate in drug delivery and biomedical areas. Natural polysaccharides in drug delivery and biomedical applications: Elsevier; 2019:59–100.
- Lei H, Xie M, Zhao Y, Zhang F, Xu Y, Xie J. Chitosan/sodium alginate modificated graphene oxide-based nanocomposite as a carrier for drug delivery. Ceram Int. 2016;42(15):17798–17805.
- Xie M, Zhang F, Liu L, et al. Surface modification of graphene oxide nanosheets by protamine sulfate/sodium alginate for anti-cancer drug delivery application. Appl Surf Sci. 2018;440:853–860.
- Ahmad MZ, Alkahtani SA, Akhter S, et al. Progress in nanotechnology-based drug carrier in designing of curcumin nanomedicines for cancer therapy: current state-of-the-art. J Drug Target.. 2016;24(4):273–293.
- Salatin S, Yari Khosroushahi A. Overviews on the cellular uptake mechanism of polysaccharide colloidal nanoparticles. J Cell Mol Med. 2017;21(9):1668–1686.
- Behzadi S, Serpooshan V, Tao W, et al. Cellular uptake of nanoparticles: journey inside the cell. Chem Soc Rev. 2017;46(14):4218–4244.
- Kumari A, Yadav SK. Cellular interactions of therapeutically delivered nanoparticles. Expert Opin Drug Deliv. 2011;8(2):141–151.
- Kou L, Sun J, Zhai Y, He Z. The endocytosis and intracellular fate of nanomedicines: Implication for rational design. Asian J Pharm Sci. 2013;8(1):1–10.
- Suryani MR, Ismail H. Preparation of curcumin nanoparticles and cellular uptake study on HeLa cells. Paper presented at International Conference on Latest Trends in Food, Biological & Ecological Sciences (Dubai:).[Google Scholar]2015.
No competing interests reported.