Introduction

Pandemic diseases have always shattered the economic and healthy stability of society. Most of the pandemics are caused by microbial infections. The occurrence of such pandemics has increased in the present-day world. The prolonged usage of synthetic antibiotics helped the infectious micro-organisms to develop resistance towards them (Natarajan, Shivakumar, & Srinivasan, 2010; Raj et al., 2013). This increased the efforts towards the production of Ayurvedic drugs from natural origin. These Ayurvedic formulations have fewer side effects and thus preferred as safe (Sayeed, Al-Bari, & Rahman, 2006).

Dashapushpam constitute a group of ten various plants namely Emilia sonchifolia (L.) DC (Dash, Abdullah, & Yahaya, 2015; Sophia, Kanniapan, Ragavendran, Raj, & Gopalakrishnan, 2012), Aerva lanata (L) Juss (Indira, 2015; Payal & Gurlaganjeet, 2015), Eclipta alba (L.) Hassk (Jaglan, Brar, & Gill, 2013; Singh, Singh, & Dwivedi, 2014), Cardiospermum halicacabum (Linn.) (Raza, 2013; Stalin, Vivekanandan, & Bhavya, 2013; Suresh et al., 2012), Biophytum sensitivum(L.) DC (Pawar & Vyawahare, 2014; Saritha & Brindha, 2015), Evolvulus alsinoides (Linn.) Linnv (Anbarasu, Selvan, Baskar, & Raja, 2016; Singh, 2008), Cynodon dactylon (Pers.) (Das, Shilpi, & Chandra, 2013; Pandey, Singh, Prasad, Singh, & Mishra, 2016), Ipomoea sepiaria Roxb. (Sayani, Ashok, & Nishteswar, 2012), Curculigo orchioides Gaertn (Irshad, Singh, Jain, & Khanuja, 2006), Vernonia cinerea L (Prabha, 2015; Varghese, Anila, Nagalekshmi, Sonu, & Resiya, 2010). They are famous for their different medicinal features such as antihelminthic, anti-diabetic, antioxidant, hepatoprotective, antidiarrheal, anti-microbial activity, anticancer, anti-inflammatory, antitumor and immune-modulatory (Bitasta & Madan, 2016; Mini et al., 2010). The different medicinal properties of these plants can be summarised as in Table 1 and Table 2.

The current work is intended to analyse the anti-bacterial property of the individual aqueous extract of the plants and their combinational formulation as the drug Dasapushpa Ghritham when tested against Bacillus cereus and Pseudomonas aeruginosa. The Ayurvedic formulation of Dasapushpa Ghritham is from “visha vaidhya jyostnika” which is an old text in Ayurveda used in ancient times. The formulation constitutes the aqueous extract of the Dashapushpam herbs as the actual content and a drug base like ghee, turmeric, sandal etc. (Krishnapriya, Sreerudran, & Hussain, 2018)

Materials and Methods

The Dashapushpam herbs were collected from different parts of Malappuram district, Kerala, India. The plants were first washed in tap water and then dried in the shade and powdered.

Extraction preparation

The Dashapushpam plants were washed in tap water and shade dried. 50g of powdered sample was weighed and taken and added to 300ml distilled water. This solution was heated for 15 min and stirred continuously. The plant extract was then cooled for 24hrs at room temperature. After that, the solution was filtered through a Whatman filter paper [no.1] and vacuum pump. The filtrate obtained was concentrated at 40^oc until all the solvents evaporated completely. Then each sample was dissolved in sterile distilled water separately (Parekh & Chanda, 2006).

Drug preparation

The Ayurvedic formulation of Dasapushpa Ghritham is taken from the Ayurvedic text Vishavaidhya Jyostnika. The method of preparation of the drug is given in the sixth chapter of the text, which deals with the viper venom and non-healing ulcers.

The content of the drug is mainly classified into two parts that are the paste of plant parts known as Kalkkam and the fresh juice of ten sacred plants known as Swarasam. The paste of Kalkam is prepared by using the various parts of plants.

Micro-organisms used

Test organisms used were Gram-negative bacteria as Pseudomonas aeroginosa, and a gram-positive bacterium was Bacillus cereus.

Anti-bacterial activity

Agar-gel diffusion technique was done to analyse the anti-bacterial property of the plant extract samples. Medium for bacterial culture was nutrient agar plates. On the 20ml solidified nutrient agar, 1ml of each bacterial suspension was inoculated by spread plate method. Each plate had 6mm diameter wells cut out in it.

The samples, positive control (Amoxicillin 30µg/ml), negative control (sterile distilled water) was added in the first, second and third well respectively. 20 µl of the plant extract (0.5 g/ml) was added into the first well of one gram-positive and one gram-negative plate as the sample. The Dasapushpa Ghritham (Ayurvedic drug) and the base(Ayurvedic drug without the plant extract) was added as the sample to the Pseudomonas aeroginosa and Bacillus cereus cultured nutrient agar plates with a positive and negative control. After inoculation, the incubation of the plates was done at 37^oc for 2-3 days. Inhibition zone was formed, and the diameter was measured around the sample well and the positive control.

Results and Discussion

The anti-bacterial assay results obtained from the agar-gel diffusion technique was given in Table 3. The Ayurvedic drug Dasapushpa Ghritham has a diameter of 1.54cm in inhibition zone with the gram-positive bacteria Bacillus cereus and 1.74cm with the gram-negative bacteria Pseudomonas aeruginosa, as shown in Figure 1. The Ayurvedic drug base without the Dashapushpam plant extracts also showed some anti-bacterial property. The Dasapushpa Ghritham base gave a 0.8cm diameter of inhibition zone with the two test organisms, as shown in Figure 2. The individual plants coming under the group of Dashapushpam shows the varying range of values for the anti-bacterial assay done with Pseudomonas aeruginosa and Bacillus cereus, which is compared in a graphical representation in Figure 3.

The positive control used was the standard drug amoxicillin which has a diameter of 1.75cm in inhibition zone with Gram-negative bacteria Pseudomonas aeruginosa and 1.85cm diameter of inhibition zone with the Gram-positive bacteria Bacillus cereus. Negative control (distilled water) used did not give a zone of inhibition. Ipomea sepiaria from the Dashapushpam family was the sample which gave the highest value for the anti-bacterial assay with a zone of inhibition of 1.85cm diameter for both the test organisms. Thus this result is equivalent to that of the standard drug (positive control) amoxicillin. Evolvulus alsinoides was the sample that gave the least value with the test organisms with the diameter of 1cm and 1.24cm each. The rest of the members of the Dashapushpam family have the anti-bacterial assay results ranging between these values.

Conclusion

After performing the anti-bacterial assay, the results prove that the members of the Dashapushpam, when used in a combination as a drug, shows significant anti-bacterial activity than when used as an individual plant extract. The Ayurvedic drug shows the anti-bacterial activity as that of the positive control antibiotic amoxicillin in the case of Pseudomonas aeruginosa. More studies are required to understand the therapeutic potential of the drug Dasapushpa Ghritham. Another important aspect of this work is that it encourages the use of a combinational formulation of the ten sacred plants (Dashapushpam) for many other therapeutic drugs. Using the plant extracts as a medical remedy has always been of great interest. The present study also proves the efficiency of using Ayurvedic herbs in medicinal formulations which can be used against disease-causing micro-organisms. A natural drug for microbial infection always has an excellent acceptance in the world of modern medicine.

Funding Support

No external funding support all the works are done by myself.

Conflict of Interest

We declare that we have no conflict of interest for this study.