Agustinus Yuswanto

Protective humoral immunity induced by surface-associated material from Actinobacillus actinomycetemcomitans in mice.

The aim of the present study was to determine the role of antibodies specific to anti-surface-associated material from Actinobacillus actinomycetemcomitans (anti-SAM-Aa) in an infection induced by this periodontopathogen in mice. When SAM-Aa obtained by saline extraction of A. actinomycetemcomitans Y4 was separated on one-dimensional gel electrophoresis, this constituent contained antigen fragments with molecular weights ranging from 14000 to 79000. Immunoblot analysis revealed that increased antigen dose/immunization resulted in increased numbers of antigen epitopes recognized by serum antibodies of the immunized mice. Rapid healing of the primary lesions and high levels of specific IgG antibodies after challenge with live A. actinomycetemcomitans were seen in the immunized mice, especially at the highest-dose level of 100 microg/immunization. Transfer of SAM-Aa-immunized, but not the SAM-Aa-immunized and adsorbed, serum prior to challenge with live bacteria led to rapid healing of the lesions in the recipient mice. Increased phagocytosis of A. actinomycetemcomitans by murine macrophages (RAW264.7 cells) was observed when this periodontopathogen was opsonized by the SAM-Aa-immunized, but not SAM-Aa-immunized and adsorbed, serum. These results suggest that in mice, SAM-Aa antigens may induce protective antibodies by acting, at least, as an opsonin against challenge with live A. actinomycetemcomitans.

Potency of Massoia Bark in Combating Immunosuppressed-related Infection.

Background: As part of our search for new potential natural resources to eradicate infection, we have revealed the prominent potency of massoia bark (Massoia aromatica Becc, Lauraceae) in combating immunosuppressed-related infection. Materials and Methods: The extract was prepared by macerating the pulverized dried bark in ethanol 95%, followed by solvent evaporation. The oil was extracted from the dried bark by steam-hydrodistillation of which preparative thin-layer chromatography was performed on the oil to isolate the active constituent, C-10 massoia lactone (ML). Anti-biofilm assay against Candida albicans was conducted on polystyrene 96 wells microtiter plates, followed by a confocal laser scanning microscope observation to get three-dimensional profiles of the affected biofilms. Effects on the hyphae development inoculated on RPMI-1640 agar plates were observed for 7 days. Influences of samples on mice macrophage phagocytosis were examined by an in vitro technique. Samples concentration tested were in the range of 2.0–0.0625 mg/mL and done in triplicate. Results: Massoia bark extracts (oil and solid phase) and ML exhibited promising activities as anti-biofilm against C. albicans at IC50 0.074% v/v, 271 μg/mL and 0.026 μg/mL, respectively. The ML did not inhibit the hyphae development at the concentration tested; however, the extracts showed inhibition at 62.5 μg/mL. Macrophage phagocytosis stimulation was correlated to the ML content. Conclusion: Massoia bark is potential to be developed as anti-infective in immunosuppressed condition of which the C10 ML (C10H16O2) plays a major role in exerting activity.

In vitro Evaluation of Massoia Bark Essential Oil and C-10 Massoialactone Potency as Immunomodulator

Abstract Massoia aromatica Becc (Lauraceae) bark essential oil has been widely used as part of traditional healing remedy in Indonesia. The major constituent, C-10 Massoia lactone has been reported previously as the antimicrobial principle. However, its effect on the immune response has not been reported so far. This research was aimed to find out the immunomodulatory effects of the Massoia essential oil and the C-10 Massoia lactone against Candida albicans as well as to explore the potential toxicities against Vero and primary culture of fibroblast cells. Material and methods: The essential oil was obtained by a steam-hydrodistillation method. The C-10 Massoia lactone was isolated from the essential oil by a preparative chromatography. Cytotoxicity assays were done on Vero and primary culture of fibroblast cells by using the MTT assay. The immunological assay were performed in vitro on lymphocytes and macrophages taken from Sprague-Dawley mice. The concentration range of samples were 30 - 2.5 μg/mL and done in triplicate. ANOVA followed by the Tukey-HSD (honestly significance diffirence) was used for the statistical analysis. All sample showed significant activation of the macrophage phagocytosis both on latex and Candida albicans in a concentration dependant manner. IC50s of the C-10 Massoia lactone and the essential oil on primary culture of fibroblast were 11.29 and 26.07 μg/ml, respectively, while on Vero cells were found at 28.35 and 37.34 μg/ml, respectively. All samples exhibited no effect on lymphocyte cell proliferation.

Effect of Massoia (Massoia aromatica Becc.) Bark on the Phagocytic Activity of Wistar Rat Macrophages

The essential oil of Massoia (Massoia aromatica Becc., Lauraceae) bark is a potential immunomodulator in vitro. This study evaluated the potential immunomodulatory effects of Massoia bark infusion on the nonspecific immune response (phagocytosis) of Wistar rats. For the in vitro assay, macrophages were treated with the freeze-dried infusion at the concentrations of 2.5, 5, 10, 20, or 40 µg/mL media. For the in vivo assay, two-month-old male Wistar rats were divided into five groups. The baseline group received distilled water at the dose of 1 mL/100 g body weight (BW), with the herbal product containing Phyllanthus niruri extract that was administered as the positive control at the dose of 0.54 mL/rat. The treatment groups received the infusion at a dose of 100, 300, or 500 mg/100 g BW. Treatments were given orally every day for 14 days. The ability of macrophage cells to phagocyte latex was determined as phagocytic index (PI), and it was observed under microscopy with 300 macrophages. The in vitro study revealed that the phagocytic activity of the infusion-treated macrophages significantly increased in comparison with that of the control macrophages in a concentration-dependent manner. Among all of the treatment concentrations, the concentration of 40 µg/mL provided the highest activity with a PI value of 70.51 ± 1.11%. The results of the in vivo assay confirmed those of the in vitro assay. The results of the present study indicate that Massoia bark can increase the phagocytic activity of rat macrophage cells.

Authentication of Wild Boar Meat in Meatball Formulation Using Differential Scanning Calorimetry and Chemometrics

An UV spectrophotometric area under curve method was developed for the estimation of Levofloxacin Hemihydrate in its mono component tablets. The spectrophotometric method for estimation employed Area under curve method for analysis using 0.1M Sodium Hydroxide as solvent for the drug Levofloxacin Hemihydrate at the wavelength range of 285-295nm. Levofloxacin Hemihydrate obeys Beer’s law in concentration range 10-50µg/ml. The recovery studies ascertained accuracy of the proposed method and the result validated according to ICH guideline. Results of analysis have been valid statistically by recovery studies. The method was successfully for evaluation of Levofloxacin Hemihydrate in tablet dosage form without the interference of common excipients.