DOI: 10.14457/tu.the.2016.1476
title: Anti-allergic activity of Trikatuk, Triphala and Trisarn remedies

description: Trikatuk remedy (TK) has long been used in Thai traditional medicine for   adaptogen and for treating disease in the rainy season. It comprises of Piper nigrum seed, Piper retrofractum flower and Zingiber officinale rhizome. Triphala remedy (TP) is used to adjust patients' elements in summer, is composed of dried fruit of three medicinal plants, namely Phyllanthus emblica, Terminalia bellirica and Terminalia chebula. Trisarn remedy (TS) is a Thai traditional medicine used in winter, consist of three plants namely Piper interruptum vine, Piper sarmentosum root and Plumbago indica root. There is no report to comparative the anti-allergic activity of three remedies and the activity related to allergy such as anti-inflammatory. Therefore, the objectives of this research were to study the anti-allergic activity of TK, TP, TS and its ingredients. Then, the remedy which show the strongest anti-allergic properties was choosed to   study anti-inflammatory effects. Three remedies were extracts boiling in water and macerated in 95% ethanol, plant ingredients were extracts with 95% ethanol to obtain 15 extracts. The extract which showed the highest anti-allergic and anti-inflammatory activities was selected to study for stability. The results of screening on antiallergy indicated that the ethanolic extract of Trikatuk remedy exhibit the highest anti-allergic activity against antigen- induced β-hexosaminidase release as a marker of degranulation in RBL-2H3 cells, with an IC50 value of 38.02±1.34 µg/ml, followed by the ethanolic extract of Piper nigrum, Piper retrofractum, Plumbago indica, Piper interruptum and Zingiber officinale (IC50 value of 44.97±6.16, 50.91±6.44, 63.55±3.77, 78.30±3.09 and 81.85±12.00 µg/ml respectively). The water extract of Trikatuk Triphala and Trisarn remedies and other plants were apparently inactive (IC50 &gt; 100 µg/ml). Thus, this research selected the ethanolic extract of Trikatuk which showed the best anti-allergic activity to continuously studied for anti-inflammatory and stability test. The results of new extract of Trikatuk remedy and its ingredients show that the ethanolic extract of Piper nigrum exhibited the highest anti-allergic activity with an IC50 value of 22.4±2.35 µg/ml. It also showed higher anti-allergic activity than Chlorpheniramine (CPM) (IC50 value 26.13±1.89 µg/ml) but not significantly different (p-value &lt; 0.05), followed by the ethanolic extract of Trikatuk with an IC50 value 28.87±1.13 µg/ml, the ethanolic extract of Piper retrofractum (IC50 value 47.49±1.03 µg/ml) and the ethanolic extract of Zingiber officinale (IC50 value 50.07±4.33 µg/ml). The ethanolic extract of Zingiber officinale and the ethanolic extract of Trikatuk exhibited the most potent anti-inflammation by inhibitory effect against nitric oxide (NO) production in RAW 264.7 cells, with an IC50 value 19.41±1.19 and 24.35±0.81 µg/ml, which was not significantly different prednisolone (IC50 value 21.93±0.37 µg/ml) (p-value &lt; 0.05), followed by the ethanolic extract of Piper nigrum and the ethanolic extract of Piper retrofractum (IC50 value 33.23±2.33 and 35.89±2.51 µg/ml, respectively). The study on chemical fingerprint was carried out using Reverse Phase High Performance Liquid Chromatography (HPLC) and including the study on specificity, linearity, limit of detection (LOD), limit of quantitation (LOQ), precision and accuracy for validate the HPLC method. The results exhibited that HPLC method showed good specificity, linearity, lower LOD and LOQ, precision and accuracy. The ethanolic extract of Trikatuk remedy was evaluated for stability under accelerated conditions (40ºC, 75±5%RH for 6 months) and evaluated for inhibitory effect of β-hexosaminidase from RBL-2H3 cells showed that Trikatuk was significantly since day 120 from day 0, and inhibition of NO production from RAW 264.7 cells showed the highly stable as the activities was not significantly different from day 0 (p-value &lt; 0.05). Piperine, 6-gingerol and 6-shogaol determined contents as marker compounds by using HPLC method. Furthermore, the amount of piperine at day 90 and day 180 were increase than day 0 (p-value &lt; 0.05) but 6-gingerol was more quickly reduced (34.57%) after day 180 and 6-shogaol content on day 15, day 30, day 60, day 90 and day 180 were not significantly different from day 0, while the amount of 6-shogaol at day 120 and day 150 were increase than day 0 and also showed significant difference (p-value &lt; 0.05). In conclusion, these findings indicated that the ethanolic extract of Trikatuk remedy showed the highest in vitro anti-allergic and anti-inflammatory activities, and active compounds of Trikatuk remedy are stable except 6-gingerol. These results can support the use of Trikatuk as a adaptogenic drug for treatment of allergic and inflammatory related diseases Trikatuk can be used instead of allergic steroid drug. The ethanolic of Trikatuk should be developed into modern medicine for anti-allergic treatment in the future. However, 6-gingerol was unstable. Thus, Trikatuk preparation should be kept in freezer for use.