Bark anatomical observations
The bark anatomical structure (Microscopic)
of P.
elongatus revealed that the thickness of the bark was found to be 500µm. The bark constitutes a deep-seated periderm, followed by a thick cortex and very hard secondary phloem. The periderm is characterized by a fairly thick, undulate and continuous segment and originated from the deeper of the cortex. Periderm involves 3-4 layers of phellem, which seems to be dark, attributable to the gathering of tannin was observed in
P.
elongatus. Consequently, phelloderm cells were found with four layers and were squarish in shape. In specific, a rectangular narrow layer, beneath the phellem was noticed. Usually, phelloderm cells are found 3-5 layers of phellem shown to be radially elongated along with dense cell organelles. The thickness of the cortex was 100µm and cortical cells showed a horizontally elongated and cylindrical shape. Moreover, a few cortical cells contain a dense accumulation of tannin was detected in
P.
elongatus bark (Fig 2).
Secondary phloem
occurs with 400µm thickness and it was considered to be the thickest region of the bark. The zone of collapsed phloem constituted and represented with dilated rays, undilated, masses of sclereids, phloem fibres and dilated parenchyma cells of phloem with collapsed sieve cells. The undilated rays have shown to be isolated uniseriate and are being radically elongated, rectangular, with numerous dilated rays thick cells. The cells were angular, oblong, isolated and randomly distributed branchy sclereids were found abundant in the collapsed phloem segment
of P.
elongatus. The fibres of phloem were varied in shape and size. Few fibres were rectangular including an elliptical lumen, the rest of the cells were small and a polygonal shape was observed (Fig 2).
Zone of non-collapsed phloem
The microscopical observation of the zone of non-collapsed phloem envisaged that it consisted of the thick radial segment covering phloem fibres and appeared in regular horizontal lines. Subsequent cells of the fibre were known to be sieve cells, which seems to be rectangular in size and shape and distinguishable based on the size. The fibres appeared 20-30 µm in the tangential plane and 15 µm in radial plane respectively, whereas sieve cells were found 40 µm in the horizontal plane and 15µm in radial plane respectively. The parenchyma cells appeared in polyhedral or squarish shape (Fig 3).
Crystal dispersement
The phloem and periderm were found, abundantly distributed with calcium oxalate crystals, which are tiny and glandular. Crystals were noticed in a thin line along the radial walls of phloem parenchyma, sieve cells and fibres obviously and they extended their appearance as thick bands, along the tangential walls in the periderm tissues (Fig 3).
Observations on the tangential longitudinal section (TLS)
On the view of the longitudinal section of
P.
elongatus, it was important to note that, phloem elements appeared to be in a vertical position as mentioned and shown to have fibres with lignified walls as depicted in the slide. Phloem rays represented as non-storied and specifically, uniseriate and heterocellular. The ray cells contain squarish cells in the middle, while margins constitute triangular cells. The range of ray cells includes 3-20 cells in height. They were measured as 60-220µm height and 25-30µm thick respectively.
The specified ray cells were not lignified. Phloem parenchyma cells have been found in narrow and vertical strands. The other cells visualized as darkly stained amorphous bodies (Sieve cells were appeared wide rather than parenchyma cells and covered profound sieve areas on the lateral side of the walls consequently, sieve cells showed oblique end walls.
The radial longitudinal section (RLS) of
P.
elongatus showed that phloem rays were found as thick ribbon or horizontal thick body. Ray cells were seen thick, cylindrical and thick-walled. It was noteworthy that no distinguishable found between procumbent and upright cells and were heterocellular. Sclerenchyma elements, fibres and sclereids were oriented vertically while parenchyma cells found in vertical strands. They contain few dark inclusions. The orsclereids have found to be large, rectangular, wedge-shaped squarish in shape (Fig 4).
Phytochemical constituents and antimicrobial activities
Phytochemical screening of the bark extract envisaged that the plant possesses tannins, flavonoids, proteins and steroids (Table 1). Negative results were detected for saponin, anthraquinones and phenols. The detected active biomolecules that occurred in the present investigation exhibited antimicrobial activity against all selected pure bacterial and fungal strains. Many studies emphasized that alkaloid, tannins and glycoside have been exhibited and they are the potential for anti-microbial actions
(Banso 2009; Gentallan et al., 2019).
However, in the present investigation, tannin and flavanoids were excess and might have been responsible for antimicrobial efficiency. Further, Recent studies have deciphered that presence of Tannins and flavonoids has been reportedly inhibiting the development of microorganisms, targeting the microbial protein to fulfil their unavailability
(Rani et al., 2017). Hence, it has been presumed that active phytochemical present in the methanol bark extract, helps to exhibit a moderate antimicrobial potential which also suggested by
Bagheri et al., 2020.
The zone of inhibition indexed the ability of active principles of the plant to elevate the activity on the concentration dependant manner as illustrated by the value of the diameter of the zone of inhibition
(Arora and Onsare, 2014). Table 2 indicated the antimicrobial activity of methanol extract of
P.
elongatus bark. The analysis revealed that methanol bark extract showed significant inhibition against
S.
aureus at the concentration of 1000µg /mL and the zone of inhibition was recorded as 13mm and 12mm and 11mm at the concentration of 1000µg/mL, 750µg/mL, 500µg/mL respectively (Fig 5). However, positive control (antibiotic) was shown to be 29µm. It was noteworthy that inhibition effects were observed relatively less against
S.
typhii, in varied concentration and not close to the effects of control, whereas against
L.
acidophillus and
V.
alginolyticus exhibited a moderate inhibition effect and rate of inhibition was maximum of 10mm was observed in both strains at 1000µg/mL.
Arora and Mahajan, 2018 screened the antimicrobial activity of aqueous extract from the bark of wild Himalayan cherry (
Prunus cerasoides) which showed the potential activity against various pathogenic microorganisms with inhibition zone ranging from 19 to 24 mm. Similarly,
S.
Albicans showed better effects rather than
Rizhopus species. It was recorded as 14, 12 and 11mm zone of inhibition against
C.
albicans with a concentration of 1000, 750 and 500 µg/mL and it exhibited 60.8% whereas
Rizhopus is shown to have possessed 58% of MIC (Table 3).
Some species of Podocarpus are used in systems of traditional medicine for fever, coughs, arthritis, sexually transmitted disease and canine distemper
(Abdillahi et al., 2011). This could be due to the presence of different phytochemical constituent and their potential activity. The bark studies also revealed the presence of calcium oxalate crystals and resin canals which are the characteristics of
Podocarpus elongatus bark.
Madivoli et al., 2018 studied the antimicrobial activity of
Prunus africana and
Harrisonia abyssinica bark and revealed that the methanolic extracts of both plants showed moderate antibacterial activity.
