A biological active artificial saliva formulation containing flower mucilage from Ceylon Spinach (Basella alba Linn.).

Ceylon Spinach (Basella albe) is an edible perennial vine found in tropical Asia and Africa, known as vegetables containing mucilage. Its mucilage from flowers was extracted by microwaving and precipitated with 95% ethanol. Five artificial saliva formulations composing of mucilage from Ceylon Spinach, calcium chloride (CaCl2), potassium chloride (KCl) and sodium fluoride (NF) were developed. The best formulation No.5 containing 0.61% of the mucilage with the non-Newtonian pseudoplastic flow (8.9 ± 0.2 cP) and the wetting time (12.50 ± 2.24 min) similar to the normal human saliva was selected. This artificial saliva formulation exhibited biological activities including an antioxidative activity by DPPH free radical scavenging with the SC50 of 14.26 ± 2.00 mg/ml (0.05 folds of ascorbic acid), and the adhesion inhibition of S. mutans on hydroxyapatite beads at 17.01 ± 7.75%, while the natural human saliva exhibited an increase bacterial adhesion of 33.10 ± 9.70%. The safety of this formulation which gave no cytotoxicity on normal human gingival fibroblasts at 99.20 ± 21.09% cell viability was also demonstrated. The results from this study have indicated high biological activity and safety of the developed formulation containing mucilage from Ceylon Spinach which is potential to be used as artificial saliva for xerostomia patients.

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 Phosphodiesterase Inhibition and Nitric Oxide Release Stimulation of Anti-Impotence Thai Medicinal Plants from "MANOSROI III" Database.

Seven plants in the top rank were selected from the "MANOSROI III" database using the two Thai keywords which meant impotence and sexual tonic. Boesenbergia rotunda (L.) Mansf. extract [EDP1-001(1)] gave the highest PDE inhibition activity of 4.36-fold sildenafil, a standard anti-impotence drug. Plumbago indica Linn. extract [EDP2-001(1)] exhibited the highest NO release stimulation activity of 666.85% which was 1.50-fold acetylcholine, a standard drug. Most selected plant extracts were nontoxic to EA.hy926 cells at 1.0 mg/mL. EDP1-001(1) exhibited the LD50 value of acute oral toxicity in male ICR mice of over 5,000 mg/kg body weight. EDP1-001(1) also indicated the improvement of sexual behaviors in the paroxetine-induced sexual dysfunction male mice with the evaluation of number of courtships (NC), mount frequency (MF), intromission frequency (IF), and ejaculatory frequency (EF) at 87.67 ± 6.17, 121.00 ± 23.50, 36.00 ± 3.21, and 13.67 ± 2.96 which were 2.63-, 1.27-, 0.53-, and 0.62-fold sildenafil-treated mice at day 14 of the treatments, respectively. The present study has not only confirmed the traditional use of Thai plants for the treatment of ED but also indicated the potential and application of the "MANOSROI III" database for Thai plant selection to be developed as ED food supplements.

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.

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.

In vitro anti-cancer activities of Job's tears (Coix lachryma-jobi Linn.) extracts on human colon adenocarcinoma.

The whole seed (W), endosperm (E) and hull (H) of five cultivars of Job's tears (Coix lachryma-jobi Linn. var. ma-yuen Stapf) including Thai Black Phayao, Thai Black Loei, Laos Black Loei, Laos White Loei and Laos Black Luang Phra Bang were processed before solvent extraction by non-cooking, roasting, boiling and steaming Each part of the Job's tears was extracted by the cold and hot process by refluxing with methanol and hexane. The total of 330 extracts included 150 methanol extracts and 180 hexane extracts were investigated for anti-proliferative activity on human colon adenocarcinoma cell line (HT-29) by the sulforhodamine B (SRB) assay. The extracts which gave high anti-proliferative activity were tested for apoptotic activity by acridine orange and ethidium bromide double staining and anti-oxidative activities including free radical scavenging and lipid peroxidation inhibition activities. The extract from the hull of Thai Black Loei roasted before extracting by hot methanol (M-HTBL-R2) showed the highest anti-proliferative activity on HT-29 with the IC50 values of 11.61 ± 0.95 µg/ml, while the extract from the non-cooked hull of Thai Black Loei by cold methanol extraction (M-HTBL-N1) gave the highest apoptosis (8.17 ± 1.18%) with no necrosis. In addition, M-HTBL-R2 and M-HTBL-N1 indicated free radical scavenging activity at the SC50 values of 0.48 ± 0.12 and 2.47 ± 1.15 mg/ml, respectively. This study has demonstrated the anti-colorectal cancer potential of the M-HTBL-R2 and M-HTBL-N1 extracts.

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.

In vitro anti-aging activities of extracts from leaves of Ma Kiang (Cleistocalyx nervosum var. paniala).

CONTEXT: Cleistocalyx nervosum (DC.) Kosterm. var. paniala (Roxb.) J. Parn. & Chantaran. (Myrtaceae) or Ma Kiang contains high amounts of phenolic compounds. Antioxidant activity of its fruit and seed has been investigated. However, limited available information concerning the biological activities of its leaves has been reported. OBJECTIVE: To investigate the in vitro anti-aging potential of young and old leaves of Ma Kiang. MATERIALS AND METHODS: Ma Kiang leaves were extracted using water, methanol, and chloroform as the solvents by cold (sonication) and hot (boiling) processes. The extracts were determined for total phenolic and flavonoid contents. The extracts (at 0.001-10 mg/ml) were tested for antioxidative and tyrosinase inhibition activities using a colorimetric method. The cytotoxicity of extracts (at 0.0001-1 mg/ml) was determined with human skin fibroblasts. Also, the extracts at 0.001, 0.01, and 0.1 mg/ml which showed no toxicity were tested for MMP-2 inhibition. RESULTS: The cold methanol extract of the old leaves showed the highest total phenolic and flavonoid contents of 511.44 ± 18.23 µg GAE/mg and 262.96 ± 2.98 µg QE/mg, respectively. This extract also gave high free radical scavenging, lipid peroxidation inhibition, and tyrosinase inhibition activities with SC50, IPC50, and IC50 values of 0.02 ± 0.004, 0.23 ± 0.13, and 0.02 ± 0.006 mg/ml, respectively. The extract at 0.1 mg/ml exhibited the highest MMP-2 inhibition of 91.14 ± 1.67%. DISCUSSION AND CONCLUSION: The anti-aging potential of the cold methanol extract from old leaves of Ma Kiang can be further developed as an anti-aging agent.

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 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.

Nanoscale gelatinase A (MMP-2) inhibition on human skin fibroblasts of Longkong (Lansium domesticum Correa) leaf extracts for anti-aging.

Leaves of Longkong which collected from Chantaburi in Thailand were extracted by the hot and cold processes using three different solvents including water, chloroform and methanol. The crude extracts were tested for antioxidative activities, tyrosinase inhibition and in vitro cytotoxicity as well as the MMP-2 inhibition activity on human skin fibroblasts for anti-aging evaluation. The hot water crude extract showed the highest antioxidative activities (DPPH radical scavenging, metal ion chelating and lipid peroxidation inhibition) with the SC50, CC50 and IPC50 values of 5.40 +/- 1.23, 32.31 +/- 0.84 and 3.29 +/- 0.30 mg/ml, respectively, and the highest tyrosinase inhibition activity with the IC50 value of 0.49 +/- 0.23 mg/ml. The extract also showed no cytotoxicity on human skin fibroblasts with the cell viability of 80.52 +/- 15.16%. It demonstrated the anti-aging potential by having the pro and active MMP-2 inhibition activity, but lower than ascorbic acid of 1.28 and 1.12 times, respectively. The semi-purified extracts were prepared from this crude extract by solvent-solvent partition. The ethyl acetate soluble fraction showed higher activities (DPPH radical scavenging, metal ion chelating and tyrosinase inhibition) than the crude extract of 23.48, 71.80 and 2.58 times, respectively. This fraction exhibited similar pro and active MMP-2 inhibitory effect to the crude extract. The results from this study have indicated the possible application of the ethyl acetate fraction of the hot water crude extract from leaves of Longkong to be developed as an anti-aging product.

Toxicity reduction and MMP-2 stimulation of papain and bromelain loaded in elastic niosomes.

The elastic niosomes (Tween 61/cholesterol/sodium cholate at 1:1:0.1 molar ratio) loaded with the protease enzymes (papain and bromelain) gave the vesicular sizes of 109.5 to 143.9 nm with the negative zeta potential of -14.7 to -30.1 mv. The elastic niosomes loaded with the standard papain (PS), extracted papain (PE), standard bromelain (BS) and extracted bromelain (BE) showed deformability index (DI values) of 1.35, 1.81, 1.22 and 1.61 times higher than their corresponding non-elastic niosomes, respectively. The elastic niosomes did not only improve the entrapment efficiency of the enzymes over the non-elastic niosomes of about 1.35 times, but also reduced the toxicity on skin human fibroblasts by SRB assay of the PS, PE, BS and BE at 1.68, 2.10, 1.56 and 1.52 times, respectively. The relative MMP-2 stimulation of PS, PE, BS and BE loaded in elastic niosomes were 1.26 +/- 0.14, 1.34 +/- 0.15, 1.09 +/- 0.09 and 1.20 +/- 0.04 for the pro MMP-2 and 1.26 +/- 0.12, 1.41 +/- 0.23, 1.01 +/- 0.08 and 1.03 +/- 0.12 for the active MMP-2, respectively in comparing to the control which were similar activity to their free enzymes. The PE loaded in elastic niosomes gave superior characteristics (low cytotoxicity and high MMP-2 stimulation) to other enzymes. The elastic niosomes can enhance the chemical stability of PE, which exhibited higher remaining contents than the free PE of 1.36 times when kept at 27 +/- 2 degrees C after 8 weeks. Therefore, the extracted papain loaded in elastic niosomes appeared to have potential to be developed as a topical product for scar treatment.