Suppression of inflammatory mediators and matrix metalloproteinase (MMP)-13 by Morus alba stem extract and oxyresveratrol in RAW 264.7 cells and C28/I2 human chondrocytes.

This study aimed to investigate the effects of Morus alba stem extract (MSE) and oxyresveratrol on the suppression of pro-inflammatory mediators in LPS-stimulated RAW 264.7 macrophages and IL-1ß-stimulated C28/I2 human chondrocyte cell line. The chondroprotective effect was also investigated using the chondrocyte cell line. First, MSE was prepared and analyzed for the amount of oxyresveratrol. The anti-inflammatory effects of MSE at various concentrations were evaluated through the inhibition of nitric oxide (NO), prostaglandin (PG)-E2 and cyclooxygenase (COX)-2 production. Oxyresveratrol at the equivalent amount found in the extract was investigated in the same manner. The chondroprotective effect was investigated through the suppression of MMP-13 production. The results showed that oxyresveratrol content in MSE was 15%. In RAW 264.7 cells, MSE (5-50 µg/mL) could inhibit the NO (24-30%) and PGE2 (11-82%) production. Oxyresveratrol at 0.75 and 7.5 µg/mL could suppress NO and also inhibited PGE2 but at only at high concentration. In the chondrocyte cell line, MSE at 5-100 µg/mL significantly decreased the PGE2 and COX-2 production by 44-93% and 17-65%, respectively. Again, oxyresveratrol at both concentrations could significantly inhibit PGE2 production by 50-92% but it inhibited COX-2 only at high concentration. In addition, MSE and oxyresveratrol was shown to significantly inhibit MMP-13 production by 14-57% and 16-56%, depending on their concentrations. The MSE demonstrates the potential to be used as an alternative treatment for reducing inflammation and preventing cartilage degradation. Its component, oxyresveratrol, may exert these effects to some extent.

In Vitro Anti-Inflammatory Activity of Morus alba L. Stem Extract in LPS-Stimulated RAW 264.7 Cells.

Morus alba L., also known as white mulberry or Mhon, has long been used in traditional medicines. This study was aimed to investigate anti-inflammatory activities of mulberry stem ethanolic extract (MSE) in lipopolysaccharide- (LPS-) stimulated RAW 264.7 macrophage cell line. The MSE was first prepared and then investigated for cell viability using the MTT assay. The anti-inflammatory activities were investigated through the inhibition of inducible nitric oxide synthase (iNOS), cyclooxygenase- (COX-) 2 mRNA expression, and iNOS protein expression using reverse transcription-polymerase chain reaction (RT-PCR) assay and immunoblotting analysis, respectively. The inhibition of nitric oxide production of the MSE was also investigated using the Griess reaction assay. The MSE concentration ranging from 10 to 40 µg/ml yielded cell viability higher than 80%. The MSE at concentrations of 20 and 40 µg/ml demonstrated anti-inflammatory activity through the inhibition of nitric oxide production via suppression of both the iNOS mRNA and protein. It was also found to inhibit the expression of COX-2 mRNA in LPS-induced RAW 264.7 cells. This study is the first to report the anti-inflammatory potential of the extract prepared from the stem of mulberry.

Morus alba L. Stem Extract Attenuates Pain and Articular Cartilage Damage in the Anterior Cruciate Ligament Transection-Induced Rat Model of Osteoarthritis.

AIM: This study was designed to investigate the anti-nociceptive effect of Morus alba stem extract as well as its cartilage protective effect in the anterior cruciate ligament transection (ACLT)-induced rat model of osteoarthritis (OA). METHODS: The anti-nociceptive effect of this plant extract was determined by measuring hind limb weight bearing, while the severity of cartilage damage to the knee joints was evaluated using the modified Mankin grading system. RESULTS: Oral administration of M. alba stem extract (56 and 560 mg/kg) significantly attenuated joint pain as indicated by a significant (p < 0.05) increase in the values of percent weight borne on the operated hind limb for the OA-induced groups that received M. alba stem extract at 56 and 560 mg/kg when compared to those of the vehicle-treated OA-induced group. In addition, a significant improvement in the Mankin score was also observed in rats treated with 560 mg/kg M. alba stem extract, which was in agreement with its pain-relieving effect. CONCLUSION: The results showed that M. alba stem extract exhibited an anti-nociceptive effect as well as cartilage protection in the ACLT-induced rat model of OA, supporting its potential use as a therapeutic treatment for OA.

Method for screening sunscreen cream formulations by determination of in vitro SPF and PA values using UV transmission spectroscopy and texture profile analysis.

Formulation of sunscreen products to obtain high values of sun protection factor (SPF) and protection from ultraviolet A (PA) is challenging work for cosmetic chemists. This study aimed to study factors affecting SPF and PA values using ultraviolet transmission spectroscopy as well as texture profiles of sunscreen formulations using 23 factorial designs. Results demonstrate that the correlation coefficient between the labeled SPF values of counter-brand sunscreen products and the in vitro SPF values was 0.901. In vitro SPF determination showed that the combination effect of phase volume ratio (PVR) and xanthan gum caused a significant increase to the SPF values of the formulations, whereas the interaction effect between PVR and stearic acid significantly decreased the SPF value. In addition, there was the interaction effect between xanthan gum and stearic acid leading to significant reduction of hardness, compressibility, and pH, but significantly increasing the adhesiveness. All tested factors did not significantly affect the cohesiveness of tested formulations. In conclusion, apart from sunscreen agents, the other ingredients also affected the SPF and PA values. The calculated SPF values range from 21 to 60. However, a selected formulation needs to be confirmed by the standard method of testing. In addition, the physical, chemical, and biological stability; shelf life; and sensory evaluation of all formulations need to be evaluated.

Methylated N-(4-N,N-dimethylaminocinnamyl) chitosan-coated electrospray OVA-loaded microparticles for oral vaccination.

The purpose of this study was to prepare microparticles entrapping ovalbumin (OVA) as a model antigen to induce immune responses in mice following oral vaccination. In this study, calcium-alginate and calcium-yam-alginate microparticles were prepared by crosslinking alginate with calcium chloride solution using an electrospraying technique. 0.1% (w/v) of methylated N-(4-N,N-dimethylaminocinnamyl) chitosan (TM65CM50CS) was used to coat microparticles entrapping an initial OVA of 20% w/w to polymer. The results indicated that the coated microparticles were spherical and had a smooth surface, with an average size of 1-3 µm, and were positively charged. In addition, the particles demonstrated a greater swelling and mucoadhesive properties than did uncoated microparticles. The in vitro release from the microparticles indicated that the coated microparticles resulted in more sustained release than uncoated microparticles. The cytotoxicity results showed that all of the formulations were safe. The in vivo oral administration demonstrated that at the same amount of 250 µg OVA, coated microparticles exhibited the highest in vivo adjuvant activity in both IgG and IgA immunogenicity.

Polymeric micelles of PEG-PE as carriers of all-trans retinoic acid for stability improvement.

The topical application of all-trans retinoic acid (ATRA) is an effective treatment for several skin disorders, including photo-aging. Unfortunately, ATRA is susceptible to light, heat, and oxidizing agents. Thus, this study aimed to investigate the ability of polymeric micelles prepared from polyethylene glycol conjugated phosphatidylethanolamine (PEG-PE) to stabilize ATRA under various storage conditions. ATRA entrapped in polymeric micelles with various PEG and PE structures was prepared. The critical micelle concentrations were 97-243 µM, depending on the structures of the PEG and PE molecules. All of the micelles had particle diameters of 6-20 nm and neutral charges. The highest entrapment efficiency (82.7%) of the tested micelles was exhibited by ATRA in PEG with a molecular weight of 750 Da conjugated to dipalmitoyl phosphatidylethanolamine (PEG(750)-DPPE) micelles. The PEG(750)-DPPE micelle could significantly retard ATRA oxidation compared to ATRA in 75% methanol/HBS solution. Up to 87% of ATRA remained in the PEG(750)-DPPE micelle solution after storage in ambient air for 28 days. This result suggests that PEG(750)-DPPE micelle can improve ATRA stability. Therefore, ATRA in PEG(750)-DPPE micelle is an interesting alternative structure for the development of cosmeceutical formulations.

Chitosan and trimethyl chitosan chloride (TMC) as adjuvants for inducing immune responses to ovalbumin in mice following nasal administration.

Chitosan of different molecular weights (Chi-P, MW=2.7x10(5) g/mol and Chi-A, MW=5.0x10(5) g/mol) and trimethyl chitosan chloride (TMC) of various degree of quaternization (DQ) including TMC-20, TMC-40 and TMC-60 were evaluated as adjuvants for inducing of immune responses to ovalbumin (OVA). OVA in solution and in alum were used as controls. Groups of BALB/c mice were immunized on days 0, 7 and 14. The IgG and IgA titers were examined on days 0, 13 and 21. It was found that for both days 13 and 21, Chi-A could elicit higher IgG responses to OVA than Chi-P. On day 13, OVA in TMC-40 induced IgG responses significantly higher than that in solution, Chi-P and TMC-60. Moreover, OVA in TMC-40 could induce IgG responses higher than OVA in alum. Although a significant difference was not observed at day 21, OVA in TMC-40 was shown to induce higher IgG responses than that in TMC-20, TMC-60 and solution. The IgA responses were the most pronounced on day 21. Again, Chi-A could elicite higher IgA responses than Chi-P and TMC-40 induced the highest IgA responses. In conclusion, these findings demonstrate that both MW of chitosan and DQ of TMC influence the level of immune induction. TMC-40 shows to be the most potent adjuvant for intranasal administration.

Targeted transfollicular delivery of artocarpin extract from Artocarpus incisus by means of microparticles.

Artocarpin (Ar), an extract of heartwood of Artocarpus incisus, possesses potent 5alpha reductase inhibitory effect. The penetration of Ar into the deeper layers of the skin where androgen receptors are present is limited. Therefore, this study was aimed to prepare alginate/chitosan (ACS) microparticles for targeted transfollicular delivery. It was found that a suitable particle size ranging from 2 to 6 microm can be prepared using the ionotropic gelation technique. Entrapment efficiency of Ar in ACS microparticles was 18.7+/-1.7%. The release of Ar from the ACS microparticles over 6 h was 0.7% of the loading dose suitable for a long-term release of Ar in the follicular ducts. The optimal growth suppression of the hamster flank organs could be achieved by topical application of Ar-ACS microparticles with a content of 0.1 mg in 5 mg microparticles to one hamster flank while the other flank (intraspecies control) showed the normal growth of the flank organs and Ar at the same concentration in solution form could not suppress the growth of the flank organs to the same extent. The efficiency of Ar 0.1 mg loaded in ACS microparticles was shown to be comparable to a dose of 1 mg Ar applied as solution. However, Ar formulated in microparticles did not show significant systemic action compared to the dermal application of an Ar solution and a flutamide preparation (1 mg) as positive control.

Effect of degree of esterification of pectin and calcium amount on drug release from pectin-based matrix tablets.

The aim of this work was to assess the effect of 2 formulation variables, the pectin type (with different degrees of esterification [DEs]) and the amount of calcium, on drug release from pectin-based matrix tablets. Pectin matrix tablets were prepared by blending indomethacin (a model drug), pectin powder, and various amounts of calcium acetate and then tableting by automatic hydraulic press machine. Differential scanning calorimetry, powder x-ray diffraction, and Fourier transformed-infrared spectroscopy studies of the compressed tablets revealed no drug-polymer interaction and the existence of drug with low crystallinity. The in-vitro release studies in phosphate buffer (United States Pharmacopeia) and tris buffer indicated that the lower the DE, the greater the time for 50% of drug release (T50). This finding is probably because of the increased binding capacity of pectin to calcium. However, when the calcium was excluded, the pectins with different DEs showed similar release pattern with insignificant difference of T50. When the amount of calcium acetate was increased from 0 to 12 mg/tablet, the drug release was significantly slower. However, a large amount of added calcium (ie, 24 mg/tablet) produced greater drug release because of the partial disintegration of tablets. The results were more pronounced in phosphate buffer, where the phosphate ions induced the precipitation of calcium phosphate. In conclusion, both pectin type and added calcium affect the drug release from the pectin-based matrix tablets.

Factors influencing the entrapment of hydrophilic compounds in nanocapsules prepared by interfacial polymerisation of water-in-oil microemulsions.

This study demonstrates the effect of drug properties and method of loading (sorption and encapsulation) on entrapment within poly(alkyl cyanoacrylate) nanocapsules prepared by interfacial polymerisation of biocompatible water-in-oil microemulsions. For small molecular weight compounds (<1000 Da), entrapment efficiency is more dependent on charge of the compound than on the method used for entrapment. Entrapment efficiency within the negatively charged nanocapsules (zeta potential approximately -30 mV) was in the order cationic compound > neutral compound > anionic compound. Only minimal differences for entrapment efficiency were noted between sorption (addition of the compound 4 h after initiation of the polymerisation) and encapsulation (addition of the compound to microemulsion prior to polymerisation). For high molecular weight compounds, the method used for entrapment however, is very important. For hydrophilic macromolecules such as proteins, high entrapment efficiencies can only be achieved by encapsulation. Entrapment of such compounds seems to be independent of the net charge of the compound being encapsulated but depended on the molecular weight. For nanocapsules prepared by interfacial polymerisation of water-in-oil microemulsions, these findings are useful as a foundation in the development of nanocapsules with desired properties.