Review of related literature These past few years, studies about the effects of certain pollutants to the anatomical

and morphological structures were made. These studies made use of different parts of the plant like leaves, stems, and petiole. The effects in the morphological structure of the plant were measured through the length of leaves, length of the petiole, width of the leaf, stem circumference, petiole circumference, thickness of leaves. On the other hand, the anatomical changes were noted through measuring the size and shape of cells, thickness of the cell walls and the orientation of the mesophyll layer both spongy and palisadein the leaves. The studies regarding the effects of air pollution to the anatomy and morphology of the plant species reviewed were conducted abroad and hopefully will be able to display the possibly same scenario here in the Philippines. The Researches A study by Assadi, Pirbalouti et al., the physiological and morphological characteristics of Eucalyptus camaldulensis Den. were observed in relation to the impact of air pollution. The experimenters randomly selected from similarly aged Eucalyptus camaldulensis trees from Gambeh park as the clean area and the Foolad industrial company of Ahvaz as the polluted area. Three replicates of fully matured leaves were collected during the summer of 2010 and were brought to the lab immediately for analysis. Right

after the cleaning of the collected leaves, the researchers proceeded to the morphological (leaf area, length of petiole, etc) characteristics measurement. As a result, the researchers found out that there have been significant lower measurements in the morphological characteristics such as the length of leaves, breadth of leaves, length of petiole, and leaf area when they were from a polluted place than when they are from the clean area. Reduction in leaf surface area and length of petiole causes less contact with environmental pollutants, specially air pollutants and improves resistance of plants against pollution (Assadi, Pirbalouti et al.). Reduction in leaf area of many plant species growing in the vicinity of heavy pollutants was observed (Seyyednejad et. al). Some hidden injury or physiological disturbance might have occurred which caused reduction in morphological and anatomical characters of plant species (Seyyednejad et. al). One of the characteristics reflecting the ability of the plant to protect against stress is the leaf length. A reduction in the leaf size and growth of aerial parts in woody plants is caused by the long-lasting impact of different pollutants and heavy metals. The previous study covered the morphological and physiological (which is not related to the current study) changes in the plants when subjected to pollution but not the anatomical changes. In a research conducted by Stevovic, Mikovilovic and CalicDragosavac, they have tried to record the changesanatomical and morphologicalmade by the plant to be

able to adapt to the polluted environment. They made use of two different cites of Africa where they have collected the Tansy. One is from the anthropogenic Ada Huja, an industrial location and the other one is from a non-anthropogenic location, Topcider Park a green area. On each cite, three different places were collected from. The plant parts that were collected were leaf, stem and root. In the anatomical study, the different leaf parts were subjected to FAA (formalin-acetic acid 10:5:85) and were dehydrated in a graded ethanol series and embedded in paraffin wax at 58C and then the sections were stained using haemotoxylin. The sections were viewed under the microscope to measure the quantitative measurements of the cuticle, mesophyll, epidermal layer, and parenchyma cells with the use of an ocular micrometer. The measurements and observations were done thrice. It was found out using the tranversal section (middle part) of the leaf that the leaves of Tansy from Ada Huja were thinner than the leaves from Topcider. It was also noted that well developed cuticle on the surface of leaves can be observed in the leaves collected from Ada Huja. The section shows that the upper and lower epidermis consists of a single layer of rectangular or orbicular cells. Epidermal cells exhibited different shapes and sizes. Multicellular trichomes were also observed on both epidermal surfaces. Thickness of mesophyll layer from Ada Huja was measured to be significantly thinner than that of the mesophyll layer of the leaves from

Topcider. Leaf mesophyll from Ada Huja consisted of 1 or 2 layers of elongated palisade cells. The palisade cells had showed a large amount of chloroplasts and large intercellular cavities. Ada Huja leaves showed thinner upper and lower palisade parenchyma than that of the Topcider plants. Surrounding the solitary vascular bundles are parenchymatous and orbicular cells in the Ada Huja plants. Also, the leaves had 2 or 3 layers of isodiametric spongy parenchymatic cells with lower intercellular activities. Upper and lower palisade parenchyma thickness from Topcider was noted as 1.3 times greater than the palisade parenchyma of Ada Huja. On the other hand, the thickness of the spongy parenchyma shows no difference between the two locations where the plants were collected. Welldeveloped vascular bundles in the leaves from both sites were observed. The cross section of the stem and root from both sites do not show specific anatomical changes. A study conducted by Gomes et al., studied the anatomical changes in Bracharia decumbens subjected to a heavy metal-containing soil. Seeds of the B. decumbens were sown to the polystyrene trays, and after they reached 10 cm in height, they were transplanted into vases containing 6L of the soil mixtures (soil was collected from a zinc mining company in Tres Marias, MG, Brazil) and then thinned. Analyses were done 120 days after subjecting the plant to the soil mixture. The anatomical analyses were done to the collected materials (roots, leaves) to FAA for 72 hours and were kept in ethyl alcohol.

A table microtome was used to obtain cross sections of the foliar mid third section and 4 +/- 0.5 cm away fro the root apex. Paradermal cuts were manually performed on the mid third of leaves. The cross sections were prepared with bleach 5% (Kraus and Arduim,1997) and an astra blue and safranine staining mixture(Kraus and Arduim, 1997). The paradermal sections were stained with a safranine solution at 1%. Glycerin 50% was used to mount the slides. All slides were examined and photographed using a Ken-a-Vision TT18 light microscope and a Canon Power Shot A620 digital camera. (Gomes et al., 2011) A software called Sigma Scan was used to determine the anatomical characteristics on three fields of five individuals per treatment. In evaluating the leaf charcacteristics, the midrib was used as a benchmark, counting the number of conducting elements of the xylem and measuring the height and width of the vascular bundle. Thickness measurements were also taken of endodermis, sclerenchyma of abaxial and adaxial surface, epidermis of adaxial and abaxial surface,, leaf blade, and bulliform cells. The area of metaxylem elements was determined by the sum of areas of three larger diameter metaxylem elements in the vascular bundle. (Gomes et al., 2011) The study used a completely randomized experimental design. The data were submitted to ANOVA, and means were compared using the Tukey test (p</= 0.05), using statistical application Sisvar 4.3. The results showed that the heavy metals absorbed by the plant caused the thickening of the abaxial and

adaxial epidermis and the larger bulliform cells found in the species can be a way to minimize water loss by transpiration, also relating to the graeter leaf blade turgor that was seen and observed in the plants subjected to contamination. Leaf curling can be a way to reduce the transpiration area on the surface, keeping stmata I a humid microclimate and therefre preventing dryness. (Turner and Jones, 1980) It was observed that exposure to to heavy metals leads to a reduction in the size of mesophyll cells(Srighar et al., 2005; Zhao et al.,2000) and the collapse of palisade ad spongy parenchyma cells (Sirdhar et al., 2005) which could support the thinned leaf blade observed in the treatments exposed to contamination. A report says that another adaptive measure to secure water flow of plants is to reduce the number of conducting elements (Baas et a., 1983). The study showed that there are really changes in the anatomical and physiological characteristics of the plants when subjected to heavy metals. Farzadfar and Zarinkamar studied the anatomical and morphological responses of Matricaria chamomilla to Cadmium and Calcium. In this experiment, seeds of chamomile was purchased and were soaked for 4 hours in distilled water and were placed on a surface of zeolite moistened with Hoagland nutrient solution and germinated 12d in the growth chamber. The seedlings were transferred into plastic containers containing 2.5 dm3 of Hoagland nutrient solution (pH 6.8). Fourteen hours (6:00 am to 8:00 pm) of photoperiod was done in growing the plants in the

growth chamber using sunlight plus metal halide lamps at an irradiance of 500 mol m-2 s-1 with a 25/20 C day/night temperature and 60-80% air humidity. Renewal of the nutrient solution was done every four days to prevent nutrient depletion. After 7 days of cultivation, Cadmium and CaCl2were added to the nutrient solution. Control plants did not receive any additional chemicals (Farzadfar and Zarinkamar, 2012). Fourteen days was the time span in exposing this species to the elements and after the 14 days, the plants were harvested and their morphological parameters were measured. In the results, anatomically, the roots showed that there is an increased size in the pericycle, epidermal, parenchyma cell and the size of intercellular spaces when subjected to CaCl2. It was also found that application of lower CaCl2 concentrations on the size of epidermal cells, pericycle, aerenchyma cells and parenchyma cells was more effective in comparison to diameter of root, central vein, phloem and xylem of roots. Anatomical root treats of chamomile seedlings decreased by the increase in Cd application, while Ca content led to alleviation of adverse effects of Cd. It was found that Cadmium acts as a repressor of the root growth and anatomical structures. Based on the results of the experiment, root as the first organ of plant in the stages of development makes use of different mechanisms in the presence of heavy metals. The researchers concluded that the interaction between Cd and Ca is useful in lessening of the Cd toxicity in chamomile seedlings. High Ca nutrition

may reduce the Cd transportation through Ca channels, leading to the lessened adverse effect of Cd on the morphology and anatomy of the plant. It was observed in the three studies regarding air pollution and effects of heavy metals to the plants show significant changes in the anatomy and morphology of the plants subjected to conditions. These changes are their adaptation to the condition to be able to survive.

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