Potent Anti-Proliferation on the Colon Cancer Cell Line (HT-29) of Liposomal Formulations Entrapped with Semi-Purified Job's Tears (Coix lacryma-jobi Linn.) Fractions.

Liposomes, entrapped with Job's Tears fractions, were prepared by the supercritical carbon dioxide fluid (scCO2) technique with and without sonication. Physical characteristics, which were the particle size, zeta potential, vesicular morphology, microviscosity and bioactivities including anti-proliferative, apoptotic and antioxidative activities of the S1L-S5L liposomal systems, were investigated. The potent anti-proliferative activity with the IC50 value of 4.44±2.31 ug/ml in a colon cancer cell (HT-29) was observed in the S5L. S5L also showed the apoptotic activity of 4.45±0.92% in an HT-29 cell. For antioxidative activities, the S3L showed the highest free radical scavenging activity and lipid peroxidation inhibition, whereas the S4L gave the highest metal chelating activity. This study has demonstrated the potent anti-proliferative activity on an HT-29 cell of the S5L liposomal formulation prepared by the scCO2 technique which can be further developed towards a novel anticancer drug.

Transfollicular enhancement of methyl myristate loaded in cationic niosomes incorporated in hair lotion.

Hair lotion containing methyl myristate loaded in cationic niosomes (HL-MMnio) composed of Brij72/cholesterol/DDAB at 7:3:0.65 molar ratio was developed. The remaining percentages of MM loaded in cationic niosomes in hair lotion were higher than free MM in hair lotion of about 1.2 times. The cumulative amounts in porcine skin and the receiver compartment of MM loaded in cationic niosomes incorporated in hair lotion were higher than those of free MM in hair lotion of 1.45 and 1.32 times, respectively. HL-MMnio showed very slightly irritation on rabbit skin, which was disappeared after 4 d. For melanogenesis induction in C57BL/6 mice with aged-induced grey body coat hairs, the highest pigmentation scores of HL-MMnio applied on the dorsal area were observed after 21 days, while hair lotion containing the free MM indicated after 35 days. This study has suggested that HL-MMnio was the high potential formulation for canities treatment.

Physicochemical properties and biological activities of Thai plant mucilages for artificial saliva preparation.

CONTEXT: Plant mucilages can be found in various parts of several Thai plants, which can be used as thickening, moisturizing, and lubricating agents in artificial saliva formulations. OBJECTIVE: The objective of this study was to evaluate the physicochemical properties, biological activity, and cytotoxicity of Thai plant mucilages. MATERIALS AND METHODS: The mucilages from Thai plants were extracted by various processes (temperature and pH variation, microwave oven, steam, and Tris-HCl buffer extraction). The viscosity and the rheology were evaluated using viscometer. Antioxidative activities including DPPH radical scavenging and metal chelating activities were investigated. The mucilages were determined for cytotoxicity on normal human gingival fibroblasts and anti-adherent activity of Streptococcus mutans. RESULTS: Mucilages from Ocimum citriodorum Vis. (Lamiaceae), Artocarpus heterophyllus Lam. (Moraceae), Abelmoschus esculentus (Linn.) Moench. (Malvaceae), and Basella alba Linn. (Basellaceae) exhibited pseudoplastic non-Newtonian rheology. The highest DPPH radical-scavenging and metal-chelating activities were observed in the mucilages from B. alba (microwave, 3 min) and A. esculentus (microwave, 1 min) with the SC50 and MC50 values (50% of scavenging activity and 50% of metal chelating activity, respectively) of 0.71 ± 0.32 and 1.11 ± 0.52 mg/ml, respectively. Most mucilages exhibited no cytotoxicity to normal human gingival fibroblasts. The mucilage from A. esculentus (microwave, 5 min) gave the shortest wetting time of 2.75 ± 0.51 min. The highest S. mutans adhesion inhibition was observed in A. esculentus (pH 11) of 5.39 ± 9.70%. DISCUSSION AND CONCLUSION: This study has indicated the suitable physicochemical and biological properties and the potential application of mucilages from Thai plants for artificial saliva preparation.

Novel application of polioviral capsid: development of a potent and prolonged oral calcitonin using polioviral binding ligand and Tat peptide.

CONTEXT: Poor absorption and proteolytic degradation are major obstacles of orally administered peptide drugs including calcitonin. Cell penetrating peptides (CPPs) and receptor binding ligands are interesting tools for the application in the delivery of these drugs. OBJECTIVE: To investigate the enhancements of in vitro and in vivo salmon calcitonin (sCT) activity by Tat, a trans-activating transcriptional peptide and VP1 peptide (V) from polioviral capsid. MATERIALS AND METHODS: Tat/sCT, V/sCT and V/Tat/sCT mixtures at various molar ratios were prepared and investigated for in vitro and in vivo activities of sCT. RESULTS: Tat could increase in vitro sCT activity both in colon adenocarcinoma (HT-29) and mouth epidermal carcinoma (KB) cells. V/sCT (6:1) showed significant increase of intracellular calcium in HT-29 cells. V/Tat/sCT (6:1:1) gave highest increase of intracellular calcium in both cells. Oral administered Tat/sCT (1:1) showed comparable hypocalcemic effect to sCT injection with prolonged action. V/Tat/sCT (6:1:1) demonstrated hypocalcemic effect at 12 h after administration but no hypocalcemic effect was observed from V/sCT. DISCUSSION: Positive charge from Tat might facilitate sCT uptake and absorption. Increasing of intracellular calcium in HT-29 cells by V but lacking of hypocalcemic effect from V/sCT in mice indicated the ligand-receptor mediated delivery of sCT by the interaction between V and PVR. CONCLUSION: Potential application of V and Tat in oral calcitonin delivery system was demonstrated. Further study in a proper PVR bearing host is still needed to provide more useful information for the application of V in the development of drug delivery systems.

Anti-proliferative and matrix metalloproteinase-2 inhibition of Longkong (Lansium domesticum) extracts on human mouth epidermal carcinoma.

CONTEXT: Longkong [Lansium domesticum Corr. (Meliaceae)] is a popular tropical plant producing economic edible fruits found mainly in Southeast Asia. However, limited information is available concerning anticancer activity of Longkong. OBJECTIVE: To investigate anticancer activities in human mouth epidermal carcinoma (KB) of Longkong extracts. MATERIALS AND METHODS: Various parts of Longkong which was collected from Northern and Eastern of Thailand were extracted by the hot and cold processes using water, chloroform, and methanol. The extracts were tested for anti-oxidative activities and anti-proliferation as well as matrix metalloproteinase inhibition on KB cells. RESULTS: The hot water extract of seeds from Northern region (NSEWH), the cold water extract of old leaves from Northern region (NOLWC), and the hot chloroform extract of young leaves from Eastern region (EYLCH) showed the highest free radical scavenging, metal ion chelating, and lipid peroxidation inhibition with SC50, MC50 and IPC50 values of 0.34 ± 0.03, 0.47 ± 1.60 and 0.86 ± 0.31 mg/ml, respectively. The hot and cold chloroform extract of young fruits from Northern region (NYFCH and NYFCC) exhibited anti-proliferation effect against KB cells with IC50 values of 603.45 ± 55.35 and 765.06 ± 46.19 mg/ml, respectively. NYFCC exhibited the highest pro- and active MMP-2 inhibition at 53.03 ± 2.65 and 31.30 ± 0.43%, more than all tested standard anticancer drugs except cisplatin. DISCUSSION AND CONCLUSION: The cold chloroform extract of young fruits from Northern region appeared to contain anticancer active compounds against KB cells because of its high anti-proliferation and MMP-2 inhibition activities.

Transdermal absorption enhancement of rice bran bioactive compounds entrapped in niosomes.

Niosomes composed of Tween 61 and cholesterol at 1:1 molar ratio were entrapped with the mixture of the three semi-purified rice (Oryza sativa L., Family Gramineae) bran bioactive compounds [ferulic acid (F), γ-oryzanol (O), and phytic acid (P)] at 0.5%, 1.5%, and 1.5%, respectively, by the supercritical CO(2) technique. The transdermal absorption by vertical Franz diffusion cells of the compounds entrapped in niosomes (Nio FOP), the unentrapped compounds (Mixed FOP), the compounds incorporated in gel and cream (Gel FOP and Cream FOP), and the compounds entrapped in niosomes and incorporated in gel and cream (Gel nio and Cream nio) was investigated. At 6 h, F and P from Nio FOP gave lower cumulative amount in viable epidermis and dermis (VED) than from Mixed FOP of 1.1 and 1.6 times, respectively, while O from Nio FOP exhibited higher cumulative amount in VED than from Mixed FOP of 2.4 times. The highest cumulative amount in VED of F, O, and P were from Gel nio, Cream nio, and Mixed FOP at 1.564 ± 0.052, 15.972 ± 0.273, and 25.857 ± 0.025 ng/cm(2), respectively. Niosomes enhanced the transdermal absorption of the hydrophobic compound O, while retarded the hydrophilic compounds F and P indicating the less systemic risk of F and P than O when entrapped in niosomes. Thus, transdermal absorption of F, O, and P appeared to depend on niosomal size, lipophilicity of the bioactive compounds, and types of formulations. These preclinical results can be applied for the design of the clinical study of the developed rice bran niosomal topical products.

Anti-aging efficacy of topical formulations containing niosomes entrapped with rice bran bioactive compounds.

CONTEXT: Rice [Oryza sativa L. (Gramineae)] bran is a rich source of phytochemicals. Its oil also contains several bioactive components that exhibit antioxidative properties such as ferulic acid (F), γ-oryzanol (O), and phytic acid (P) which can be a new source of cosmetic raw materials. OBJECTIVE: To evaluate the anti-aging effects of the gel and cream containing niosomes entrapped with the rice bran bioactive compounds. MATERIALS AND METHODS: The semi-purified rice bran extracts containing F, O, and P which indicated the growth stimulation of human fibroblasts and the inhibition of MMP-2 by sulforhodamine B and gelatin zymography, respectively, were entrapped in niosomes by supercritical carbon dioxide fluid (scCO(2)) and incorporated in gel and cream formulations. The skin hydration, elasticity, thickness and roughness, and pigmentation in human volunteers after treated with these gel and creams were investigated by corneometer, cutometer, visiometer, and mexameter, respectively. RESULTS: Gel and cream containing the semi-purified rice bran extracts entrapped in niosomes gave no sign of erythema and edema detected within 72 h on the shaved rabbit skin by the closed patch test investigated by mexameter and visual observation, respectively. These formulations also demonstrated higher hydration enhancement and improvement of skin lightening, thickness, roughness, and elasticity on the skin of 30 human volunteers within the 28-day treatment not more than 9, 27, 7, 3, and 3 times, respectively. DISCUSSION AND CONCLUSIONS: The formulations containing niosomes entrapped with the rice bran bioactive compounds gave superior clinical anti-aging activity which can be applied as a novel skin product.

Antioxidant activities and skin hydration effects of rice bran bioactive compounds entrapped in niosomes.

Bioactive compounds [ferulic acid (F), gamma-oryzanol (O) and phytic acid (P)] in rice bran have been widely used as antioxidants in skin care products. However, one of the major problems of antioxidants is the deterioration of their activities during long exposure to air and light. Niosomes have been used to entrap many degradable active agents not only for stability improvement, but also for increasing skin hydration. The objective of this study was to determine antioxidant activities [by in vitro ORAC (oxygen radical absorbance capacity) and ex vivo lipid peroxidation inhibition assay] and in vivo human skin hydration effects of gel and cream containing the rice bran extracts entrapped in niosomes. Gel and cream containing the rice bran extracts entrapped in niosomes showed higher antioxidant activity (ORAC value) at 20-28 micromol of Trolox equivalents (TE) per gram of the sample than the placebo gel and cream which gave 16-18 micromolTE/g. Human sebum treated with these formulations showed more lipid peroxidation inhibition activity than with no treatment of about 1.5 times. The three different independent techniques including corneometer, vapometer and confocal Raman microspectroscopy (CRM) indicated the same trend in human skin hydration enhancement of the gel or cream formulations containing the rice bran extracts entrapped in niosomes of about 20, 3 and 30%, respectively. This study has demonstrated the antioxidant activities and skin hydration enhancement of the rice bran bioactive compounds when entrapped in niosomes and incorporated in cream formulations.

In vitro and in vivo skin anti-aging evaluation of gel containing niosomes loaded with a semi-purified fraction containing gallic acid from Terminalia chebula galls.

CONTEXT: The galls of Terminalia chebula Retz. (Combretaceae) frequently appear in many Thai Lanna medicinal plant recipes for promotion of longevity. OBJECTIVE: The objective of this study was to evaluate the skin anti-aging of gel containing niosomes loaded with a semi-purified fraction containing gallic acid from T. chebula galls. METHOD: The semi-purified fraction containing phenolic compounds including gallic acid isolated from T. chebula galls loaded in non-elastic or elastic niosomes, and its developed gel, were evaluated for rabbit skin irritation by the closed patch test and skin anti-aging in human volunteers by measuring skin elasticity and roughness. RESULTS: Gel containing the fraction unloaded (SS) or loaded in non-elastic (SN) or elastic (SE) niosomes and gallic acid loaded in non-elastic (GN) or elastic (GE) niosomes showed no skin irritation, whereas the unloaded gallic acid (GS) gave the irritation in rabbit's skin by the closed patch test. The % parameter changes of skin elastic recovery and skin elastic extension when applied with SN and SE gels were +28.73 and +32.57; -21.25 and -22.63%, respectively. SN and SE gel also showed a significant decrease of the maximum and average roughness values with the parameter changes of -29.43 and -32.38; -39.47 and -35.28%, respectively. CONCLUSION: The semi-purified fraction loaded in niosomes indicated not only higher chemical stability of gallic acid containing in the fraction, but also more in vivo anti-aging activities than the unloaded fraction when incorporated in gel.

Transdermal absorption enhancement of gel containing elastic niosomes loaded with gallic acid from Terminalia chebula galls.

OBJECTIVES: To investigate transdermal absorption enhancement of gel containing elastic niosomes loaded with gallic acid in the semipurified fraction isolated from Terminalia chebula Retz. (Combretaceae) galls. MATERIALS AND METHODS: Nonelastic and elastic niosomes loaded with gallic acid in pure form or in the semipurified fraction were developed. Rat skin permeation by vertical Franz diffusion cells of gallic acid from various gel formulations containing elastic niosomes loaded with gallic acid or the semipurified fraction was performed. RESULTS: Elastic and nonelastic niosomes loaded with gallic acid or the semipurified fraction exhibited the mixture of unilamellar and multilamellar structures with negative zeta potential values and in the size range of 200-400 nm. Both loaded elastic and nonelastic niosomes showed good physical and chemical stability for 3 months. The percentages remaining of gallic acid in nonelastic were slightly higher than in elastic niosomes. Both elastic and nonelastic niosomes retarded rat skin permeation of the loaded pure gallic acid, while enhanced the loaded gallic acid in the semipurified fraction. However, elastic niosomes exhibited higher percentages of gallic acid through rat skin than the nonelastic niosomes. DISCUSSION AND CONCLUSION: This study has demonstrated the potential of niosomes, especially elastic niosomes, for the enhancement of chemical stability and rat skin transdermal absorption of gallic acid in the semipurified fraction from T. chebula galls, which will be beneficial for topical antiaging application.

Entrapment enhancement of peptide drugs in niosomes.

The objective of this study was to enhance the entrapment of various charged peptide drugs [(bacitracin (BCT), insulin and bovine serum albumin (BSA)] in niosomes by modifying the vesicular charge compositions. Cationic, anionic and neutral niosomes were prepared from sorbitan monostearate (Span 60) or polyoxyethylene sorbitan monostearate (Tween 61), cholesterol (CHL), dimethyldioctadecylammonium bromide (DDAB) and/or dicetyl phosphate (DP) in distilled water, by freeze dried empty liposome (FDEL) method. Morphology and vesicular sizes of the vesicles were investigated by optical microscope, TEM, X-ray diffractometry and dynamic light scattering. The entrapment efficiency of the peptides in niosomes was determined by gel electrophoresis and gel documentation. After reconstitution of the empty niosomal powder in phosphate buffer pH 7.0 containing the peptide drugs, they were oligolamellar membrane structure, with the sizes of 40-60 nm, except the neutral niosomes entrapped with insulin and cationic niosomes entrapped with BSA which showed the sizes of 0.1-1.3 microm and 100-150 nm, respectively. The zeta potential values of neutral, cationic and anionic niosomes entrapped with BSA, insulin and BCT were -22.3 +/- 1.52, -30.7 +/- 2.92 and +22.68+/- 1.31 mV, respectively. The entrapment efficiency of BSA, BCT and insulin in neutral niosomes (Tween 61/CHL at 1 : 1 molar ratio) was 72.94, 69.89 and 10.26%, in cationic niosomes (Tween 61/CHL/DDAB at 1 : 1 : 0.05 molar ratio) was 84.54, 32.85 and 87.15% and in anionic niosomes (Tween 61/CHL/DP at 1 : 1 : 0.05 molar ratio) was 50.13, 90.88 and 44.31%, respectively. The highest entrapment efficiency of BSA, BCT and insulin at 72.94, 90.88 and 87.15 was observed in neutral, anionic and cationic niosomes, respectively. The results from this study has suggested the appropriate niosomal formulation to entrap the peptides with different charges and polarity for pharmaceutical application.

Potent in vitro Anti-mouth Cancer (KB) and Immunostimulating Activities of the Job's Tears (Coix lachryma-jobi Linn.) Seed Semi-purified Extract Cocktails Containing Linoleic Acid.

The crude methanolic and hexane extracts of non-cooked, steamed and roasted from three Job's Tears cultivars were prepared and further semi-purified by liquid-liquid extraction techniques and silica gel column. The six single semi-purified extracts (F1-F6) were combined as nine cocktails (CT1, CT6, CT8, CT13, CT14, CT21, CT24, CT25 and CT31) according to the IC50 values from the preliminary study and investigated for anti-proliferative and apoptotic induction on mouth cancer cell line (KB) and immunostimulating as well as antioxidative activities. The highest anti-proliferative activity was observed in CT13 showing the IC50 value of 0.53±0.45 µg/mL which was higher than 5-fluorouracil and doxorubicin of 20.34 and 1.60 times, respectively. CT1 which was the combination of F1-F6 and CT13 which was the combination of F4-F6 exhibited significant strong synergistic activity with the combination index value (CI) of 0.28. CT1 at 200 µg/mL showed the highest percentages of apoptotic cells (40.65±10.97%) with no necrotic cells, but lower than cisplatin (100 µg/mL) of 2.18 times. CT14 gave the highest immunostimulating activity with the phagocytosis percentage of 13.0±1.7%, but lower than lipopolysaccharide of 1.08 times. CT31 gave the highest free radical scavenging and metal chelating activities with the SC50 and MC50 values of 0.73±0.07 and 1.99±0.24 µg/mL, but lower than ascorbic acid and EDTA of 18.25 and 4.33 times, respectively. The linoleic acid contents related to anti-cancer activity were also examined by HPLC. This study has demonstrated that CT1 composing of F1-F6 at the percentage ratio of 0.71:2.06:81.38: 8.47:4.92:2.46 was the potential cocktails of the semi-purified extracts from the Job's Tears which can be further developed as a novel active compound for oral cancer treatment.

Potent in vivo anticancer activity and stability of liposomes encapsulated with semi-purified Job's tear (Coix lacryma-jobi Linn.) extracts on human colon adenocarcinoma (HT-29) xenografted mice.

The in vivo anticancer activity and stability of liposomes encapsulated with semi-purified Job's tear (Coix lacryma-jobi Linn.) extracts (S5L), prepared by supercritical carbon dioxide fluid technique, on human colon adenocarcinoma (HT29) xenografted mice were investigated. For the stability and the physicochemical characteristics, S5L showed a high stability of pH, good dispersibility, small particle size and stable zeta potential. Liposomes can protect linoleic acid in the extract comparing with the free S5. S5L kept at 4 °C for 3 months showed the highest linoleic acid content of 63.50%, whereas at 45 °C, the lowest linoleic acid content of 42.66% was observed. The anticancer activity and toxicity on xenografted mice were observed for 14 days. At the end of the experiment, the relative tumor volume (RTV) in the S5L-treated xenografted mice showed a significant RTV reduction. The high dose of S5 and S5L were potent with the highest inhibition of tumor growth of 48.67 and 54.75%, which was 86.94% and 97.81% of 5-fluorouracil, respectively. The apoptotic activity was shown in xenografted mice treated with S5 at medium and high dose, S5L, 5-fluorouracil and commercial product. All treated xenografted mice showed no toxic signs and symptoms, abnormality of internal organs histopathology and blood chemistry. This study has demonstrated the high physicochemical stability of liposomes encapsulated with semi-purified Job's tear extract and their potent anticancer activity on human colon adenocarcinoma xenografted model with the potential for further development to anticolon cancer drug.

Potent in vitro anti-proliferative, apoptotic and anti-oxidative activities of semi-purified Job's tears (Coix lachryma-jobi Linn.) extracts from different preparation methods on 5 human cancer cell lines.

ETHONOPHARMACOLOGICAL RELEVANCES: Job's tears (Coix lacryma-jobi Linn.) is commonly used as an herbal medicine. The low incidence of cancer has been observed in the area of China where regularly consume Job's tears. The effects of preparation methods of Job's tears on anti-cancer activities were investigated. Potent in vitro anti-proliferative, apoptotic and anti-oxidative activities of semi-purified Job's tears extracts from different preparation methods on 5 human cancer cell lines comparing with standards and commercial product were observed. AIM OF STUDY: To study the anti-proliferative, apoptotic and anti-oxidative activities of semi-purified Job's tear extracts from different preparation methods on 5 human cancer cell lines. MATERIALS AND METHODS: The crude methanolic extracts of non-cooked, steamed and roasted Job's tears cultivars were prepared and further semi-purified by liquid-liquid extraction techniques. Both of crude and semi-purified extracts were tested for anti-proliferative, apoptotic induction, anti-oxidative activities and phytochemicals content. RESULTS: The highest yields of crude and-purified extracts were 4.60% and 1.46%, respectively. In crude extracts, the steamed whole Thai Black Loei Job's tears (W-TBL-S1) extract showed the highest anti-proliferative activity in mouth epidermal carcinoma cell (KB) at the IC50 of 43.61±0.76µg/ml (0.005 folds of doxorubicin), whereas the roasted whole Laos White Loei Job's tears (W-LWL-R2) extract showed the highest apoptotic activity in cervical adenocarcinoma (HeLa) at 21.52±1.50% (0.22 and 15.05 folds of doxorubicin and commercial product, respectively). After liquid-liquid extraction, almost all of the semi-purified extracts showed increases in anti-proliferative activity. Ethyl acetate fraction of the roasted whole Laos White Loei Job's tears (W-LWL-R2) showed the highest anti-proliferative activity in HeLa cell at the IC50 of 0.97±0.82µg/ml (7.82 and 45.39 folds of doxorubicin and crude extract, respectively) and apoptotic activity of 18.77±6.31% (0.19 folds of doxorubicin). The commercial product showed no anti-proliferative activity in all cell lines but induced apoptosis in HeLa cell at 1.43±0.34%. The butanol and hexane soluble fractions of the roasted whole of Laos White Loei Job's tears (W-LWL-R2) showed the highest free radical scavenging (SC50) and metal chelating activity (MC50) of 0.31±0.06mg/ml (0.64 folds of ascorbic acid) and 0.08±0.01mg/ml (6.37 folds of EDTA), respectively. All ethyl acetate fractions contain high content of carotenoid and tannin, whereas the hexane soluble fraction of the roasted hull of Laos Black Loei Job's tears (H-LBL-R1) showed the highest linoleic acid content of 8.09±0.74% w/w. CONCLUSIONS: This study has demonstrated the potent anti-cancer activity of the semi-purified extracts from roasted Job's tears with high potential for further development to modern anti-cancer drug.

Hypoglycemic activity and stability enhancement of human insulin-tat mixture loaded in elastic anionic niosomes.

This study aimed to investigate the synergistic effect of trans-activator of transcription (Tat) and niosomes for the improvement of hypoglycemic activity of orally delivered human insulin. The elastic anionic niosomes composing of Tween 61/cholesterol/dicetyl phosphate/sodium cholate at 1:1:0.05:0.02 molar ratio loaded with insulin-Tat mixture (1:3 molar ratio) was prepared. Deformability of the elastic anionic niosomes decreased after loaded with the mixture of 1.35 times. For the in vitro release, the insulin (T10 = 4 h) loaded in the elastic anionic niosomes indicated the slower release rate than insulin in the mixture (T10 = 3 h) loaded in niosomes. At room temperature (30 ± 2 °C), the mixture loaded in elastic anionic niosomes was more chemical stable than the free mixture of 1.3, 1.4 and 1.7 times after stored for 4, 8 and 12 weeks, respectively. Oral administration in the alloxan-induced diabetic mice of the mixture loaded in elastic anionic niosomes with the insulin doses at 25, 50 and 100 IU/kg body weight indicated significant hypoglycemic activity with the percentage fasting blood glucose reduction of 1.95, 2.10 and 2.10 folds of the subcutaneous insulin injection at 12 h, respectively. This study has demonstrated the synergistic benefits of Tat and elastic anionic niosomes for improving the hypoglycemic activity of the orally delivered human insulin as well as the stability enhancement of human insulin when stored at high temperature. The results from this study can be further developed as an effective oral insulin delivery.

In vitro anti-proliferative activity on colon cancer cell line (HT-29) of Thai medicinal plants selected from Thai/Lanna medicinal plant recipe database "MANOSROI III".

ETHNOPHARMACOLOGICAL RELEVANCE: Thai/Lanna region has its own folklore wisdoms including the traditional medicinal plant recipes. Thai/Lanna medicinal plant recipe database "MANOSROI III" has been developed by Prof. Dr. Jiradej Manosroi. It consists of over 200,000 recipes for all diseases including cancer. To investigate the anti-proliferative and apoptotic activities on human colon cancer cell line (HT-29) as well as the cancer cell selectivity of the methanolic extracts (MEs) and fractions of the 23 selected plants from the "MANOSROI III" database. MATERIALS AND METHODS: The 23 selected plants were extracted with methanol under reflux and evaluated for their anti-proliferative activity by sulforhodamine B assay. The 5 plants (Gloriosa superba, Caesalpinia sappan, Fibraurea tinctoria, Ventilago denticulata and Psophocarpus tetragonolobus) with potent anti-proliferative activity were fractionated by liquid-liquid partition to give 4 fractions including each hexane (HF), methanol-water (MF), n-butanol (BF) and water (WF) fractions. They were tested for anti-proliferative activity and cancer cell selectivity. The ME and fractions of G. superba which showed potent anti-proliferative activity were further examined for morphological changes and apoptotic activities by acridine orange (AO)/ethidium bromide (EB) staining. RESULTS: The ME of G. superba root showed active with the highest anti-proliferative activity at 9.17 and 1.58 folds of cisplatin and doxorubicin, respectively. After liquid-liquid partition, HF of V. denticulata, MFs of F. tinctoria, V. denticulata and BF of P. tetragonolobus showed higher anti-proliferative activities than their MEs. The MF of G. superba indicated the highest anti-proliferative activity at 7.73 and 1.34 folds of cisplatin and doxorubicin, respectively, but only 0.86 fold of its ME. The ME and HF, MF and BF of G. superba and MF of F. tinctoria demonstrated high cancer cell selectivity. At 50 µg/ml, ME, HF, MF and BF of G. superba demonstrated higher apoptotic activities than the two standard drugs. CONCLUSIONS: This present study has not only confirmed the traditional use of the Thai/Lanna medicinal plant recipes for cancer treatments, but also the potential of the selected plant, G. superba for the further development as a modern anti-cancer drug.

Potent Antiproliferative Effect on Liver Cancer of Medicinal Plants Selected from the Thai/Lanna Medicinal Plant Recipe Database "MANOSROI III".

Thai/Lanna medicinal plant recipes have been used for the treatment of several diseases including liver cancer. In this study, methanolic extracts (MEs) of 23 plants were tested for antiproliferative activity on human hepatoma cell line (Hep G2) by the sulforhodamine B (SRB) assay. Nine MEs with potent antiproliferative activity (IC50 < 100 µg/mL) were obtained and further semipurified by liquid/liquid partition extraction. The semipurified fractions were tested for the antiproliferative and antioxidative activities. ME of Stemona collinsae and the semipurified extract and methanol-water fraction (MF) of Gloriosa superba gave the highest antiproliferative activity on HepG2 which were 4.79- and 50.07-fold cisplatin, respectively. The semipurified fractions showed an increased antiproliferative activity. MF of Caesalpinia sappan and HF of Senna alata showed the highest free radical scavenging and metal chelating activities, respectively. The compound in n-hexane fraction (HF) of Ventilago denticulata which showed an increase in antiproliferative activity comparing to its ME was isolated and identified as emodin. This study has demonstrated the potential of the ME from S. collinsae, MF from G. superba, and emodin isolated from V. denticulata, for further development as an antiliver cancer agent.

Transdermal absorption enhancement of papain loaded in elastic niosomes incorporated in gel for scar treatment.

Papain is one of the protease enzymes from Carica papaya latex which is widely used in dermatology for scar treatment. The aim of this study was to compare the penetration of papain from gel formulations containing niosomes and nanospheres loaded with papain. The vesicular sizes of all niosomes and nanospheres in the gel formulations were in the range of 220.7-520.2 nm. Papain loaded in elastic niosomes and incorporated in gel exhibited the accumulate amounts and fluxes of 0.226 mg/cm² and 0.029 mg/cm²/h in the whole rat skin and 0.220 mg/cm² and 0.037 mg/cm²/h in the receiving solution, which were 3.10, 2.38 and 2.24, 2.25; 10.08, 7.78 and 4.92, 4.93; 4.86, 3.71 and 7.38, 7.38 times more than that from gel containing papain loaded in non-elastic niosomes, PLGA nanospheres and in solution, respectively, investigated by Franz diffusion cells at 6h. All gel formulations incorporated with papain loaded in niosomes and nanospheres gave no irritation on rabbit skin. Gel containing papain loaded in elastic niosomes gave superior chemical stability to gel containing free papain of 1.13, 1.29 and 1.35 times when stored at 4 ± 2, 27 ± 2 and 45 ± 2°C after 3 months, respectively. After 28 days of application, gel containing papain loaded in elastic niosomes (GEN) exhibited higher reduction of hypertrophic scars of the induced scar on rabbits' ears determined by a vernier caliper than gel base (GB), gel containing free papain (GS), and gel containing papain loaded in non-elastic niosomes (GNN) of 10.20, 2.73 and 2.31 times, respectively. For histological examination, the numbers of collagen fibres and the height of the scars treated with GEN were significantly decreased compared with the control group. This study has demonstrated the potential of niosomes, especially the elastic niosomes, for the enhancement of rat skin transdermal absorption of papain and the improvement of scar reduction in rabbit ear model which will be beneficial for the development of topical products for scar treatment.

Melanogenesis of methyl myristate loaded niosomes in B16F10 melanoma cells.

The objective of this study was to compare the charge effect of methyl myristate loaded in neutral (Brij 72/cholesterol at 7:3), cationic (Brij 72/cholesterol/dimethyl dioctadecyl ammonium bromide at 7:3:0.65) and anionic niosomes (Brij 72/cholesterol/dicetyl phosphate at 7:3:0.65) for physicochemical characteristics, cytotoxicity in fibroblasts and B16F10 melanoma cells as well as melanogenesis induction activity. The maximum loading and percentage entrapment of methyl myristate were 4.5, 90.68 +/- 7.95 in neutral; 11.0, 92.54 +/- 6.32 in cationic and 0.1% w/w, 74.43 +/- 1.86% in anionic niosomes, respectively. All methyl myristate loaded niosomes were in unilamellar structure under transmission electron microscope and in nanosize at initial and after 3-month storage. The percentages of methyl myristate remaining in all niosomes kept at 4 +/- 2, 30 +/- 2 and 45 +/- 2 degrees C for 3 months were about 82, 74 and 72%, respectively, while the dry free methyl myristate indicated 97.82 +/- 1.74, 96.56 +/- 2.91 and 91.39 +/- 4.32%, respectively. Blank neutral, blank cationic and methyl myristate loaded neutral and cationic niosomes exhibited moderate cytotoxicity in fibroblasts and B16F10 melanoma cells at 56.64 +/- 3.19, 59.72 +/- 1.51; 73.81 +/- 2.86, 82.51 +/- 0.20; 47.34 +/- 2.13, 52.67 +/-2.78 and 73.20 +/- 3.49, 84.34 +/- 2.75% cell viability, respectively. Blank anionic and methyl myristate loaded anionic niosomes indicated no cytotoxicity in both cells. Cytotoxic ratio of cell viability in normal and cancer cells of all niosomes indicated no toxic effect to normal cells. Methyl myristate loaded cationic niosomes demonstrated the highest melanin induction with tyrosinase activity of 1.42 and 1.70 folds of the control and 1.14 and 1.59 folds higher than theophylline, respectively. This study has suggested the potential of methyl myristate loaded cationic niosomes for canities treatment.

Potent enhancement of transdermal absorption and stability of human tyrosinase plasmid (pAH7/Tyr) by Tat peptide and an entrapment in elastic cationic niosomes.

Enhancement of transdermal absorption through rat skin and stability of the human tyrosinase plasmid (P) using Tat (T) and an entrapment in elastic cationic niosomes (E) were described. E (Tween61:cholesterol:DDAB at 1:1:0.5 molar ratio) were prepared by the freeze-dried empty liposomes (FDELs) method using 25% ethanol. TP was prepared by a simple mixing method. TPE was prepared by loading T and P in E at the T:P:E ratio of 0.5:1:160 w/w/w. For gel formulations, P, TP, PE and TPE were incorporated into Carbopol 980 gel (30 µg of plasmid per 1 g of gel). For the transdermal absorption studies, the highest cumulative amounts and fluxes of the plasmid in viable epidermis and dermis (VED) were observed from the TPE of 0.31 ± 0.04 µg/cm(2) and 1.86 ± 0.24 µg/cm(2)/h (TPE solution); and 4.29 ± 0.40 µg/cm(2) and 25.73 ± 2.40 µg/cm(2)/h (TPE gel), respectively. Only plasmid from the PE and TPE could be found in the receiving solution with the cumulative amounts and fluxes at 6 h of 0.07 ± 0.01 µg/cm(2) and 0.40 ± 0.08 µg/cm(2)/h (PE solution); 0.10 ± 0.01 µg/cm(2) and 0.60 ± 0.06 µg/cm(2)/h (TPE solution); 0.88 ± 0.03 µg/cm(2) and 5.30 ± 0.15 µg/cm(2)/h (PE gel); and 1.02 ± 0.05 µg/cm(2) and 6.13 ± 0.28 µg/cm(2)/h (TPE gel), respectively. In stability studies, the plasmid still remained at 4 ± 2 °C and 25 ± 2 °C of about 48.00-65.20% and 27.40-51.10% (solution); and 12.34-38.31% and 8.63-36.10% (gel), respectively, whereas at 45 ± 2 °C, almost all the plasmid was degraded. These studies indicated the high potential application of Tat and an entrapment in elastic cationic niosomes for the development of transdermal gene delivery system.