Combined effects of NPK fertilizer with foliar application of benzyladenine or gibberellic acid on Dracaena marginata ‘Bicolor’ grown in different potting media

A pot experiments was carried out to evaluate the influence of growing media and combined treatments of NPK with either benzyladenine or gibberellic acid on growth, chemical constituents and anatomical structure of Dracaena marginata ‘Bicolor’. The plants were grown in two growing media; peat-moss, peat-moss+ sand (1:1, v v-1), received monthly NPK fertilizers (2 and 4 g pot-1) combined with either of benzyl adenine (BA) at 100 and 150 ppm or gibberellic acid (GA3) at 150 and 250 ppm, while the control plants received no treatments. As general, the results indicated that, peatmoss was superior to peat-moss+ sand medium on increasing most of vegetative growth parameters in terms of plant height, number of leaves/plant, leaf area, root length, as well as fresh and dry weights of leaves, stems and roots/plant, besides some macro elements represented in N, P, K, Ca and Mg% in both leaves and stems. While plants grown in peat-moss+ sand possessed significantly higher contents of total chlorophylls, total carbohydrates, Cu, Fe, Mn, Zn and B than those grown in peatmoss alone. Plants received combined NPK with either BA or GA3 resulted in significant increases in most of morphological and chemicals content over the control plants and it was outstanding that, GA3 was more effective than BA when they were combined with NPK. It can be concluded that for the highest quality, quantity growth and economic production of Dracaena marginata ‘Bicolor’, the plants could be grown in a medium of peat-moss and supplied monthly with NPK fertilizer at 2 g plant-1 along with foliar sprayed with 250 ppm GA3.


Introduction
The genus Dracaena comprising about 40 species belongs to the family Ruscaceae, only six species are cultivated and used as foliage plants including D. deremensis, D. fragrans, D. reflex, D. sanderiana, D. marginata and D. surculosa. Dracaena marginata known as Madagascar dragon is evergreen ornamental shrub or small tree. It is growing to 3-5 m tall with a spread of 0.9-1.2 m. The leaves are slender, sword-shaped, slightly flexible having pleasant appearance (Huxley, 1992;Odenwald and Turner, 2006).
Growing media plays a vital role in plant growth and development of indoor potted plants. The physical and chemical properties of used media are the dominant factors affecting plant growth; hence these properties influence availability of nutrients and water for the plant growth and penetration of its roots in the soil. Peat moss is the most widely organic substrate used for the preparation of growing media for the potted ornamental plants. However, it is very expensive so that its replacement with cheaper available substrates is necessary (Shahid et al., 2017). Sand is generally the least expensive of inorganic amendments added to growing media for achieving and maintaining a structural system of macro pores that improve aeration and drainage. It was found that peatmoss alone increased the growth of different indoor plants (Mousa et al., 2015;Younis et al., 2016;Badran et al., 2017), while other studies demonstrated that peat + sand (2:1) medium increased the growth and chemical composition parameters of Celosia argentea (Abd el Gayed and Attia, 2018).
Fertilizers are essential for production of healthy potted ornamental plants. The NPK fertilization has a positive effect on the various biochemical processes that occur within the plant giving normal plant growth and development. Different NPK fertilization treatments have favorably influenced on growth of several foliage plants (El-Naggar and Ahmad, 2016;Abou Dahab et al., 2017;El-Sayed and Ismail, 2017;Mohamed, 2018).
Concerning Cytokinins are a large group of plant hormones which play a pivotal role as active molecules in many physiological processes of plant growth and development. Benzyl adenine (BA) had been newly used to maintain or increase the quality of different ornamental plants (Askari-Khorasgani and Mortazaeinezhad 2016; Matak et al., 2017), Moreover, its ability to increase number of chloroplasts, chlorophyll synthesis, bud differentiation and branching had been reported (Kochhar and Sukhbir, 2020). It was reported that its application induced vegetative growth, photosynthesis rate, soluble sugars, indoles, soluble phenols and the contents of N, P and K in organs of croton plants (Ibrahim et al., 2010). The leaf area and fresh weight of leaves and stems had been recorded (Fuadi et al., 2014).
Gibberellins (GAs) can influence different biological processes and plant growth, i.e. from vegetative growth to flowering, and also stimulate aspects of flower development. In addition, gibberellins effect on other developmental responses such as dormancy, sex expression, leaf and fruit senescence, root and fruit growth and development of seeds in the fruit (Plackett and Wilson, 2016;Satish and Manju, 2018). The beneficial effect of GA 3 on different indoor plants had been reported by prior research who concluded that GA 3 induced flowering (Huang et al., 2015;Gad et al., 2016) promoted plant height, leaf area, number of leaves, stem diameter, fresh and dry weights, relative growth rate as well as increased the contents of total chlorophylls, total carbohydrates, and macronutrients (Mazher et al., 2014;Hananfy et al., 2019).
Furthermore, previous studies had been conducted in order to evaluate the interactive effect of BA with GA 3 (Henschke et al., 2015;Gabrel et al., 2018;Sardoei, 2018), NPK fertilizer with BA (Ngapui et al., 2018), NPK with GA 3 (El-Sayed et al., 2016) and NPK with potting media (Badran et al., 2017;Mohamed, 2018). However, there is lake of researches about the influence of potting media and NPK along with BA or GA 3 as growth regulators on the indoor plants and their anatomical structure. Therefore, the main object of present study was to determine the effect of different growing media and combined NPK with either BA or GA 3 and their interaction effects on the growth, chemical constituents and anatomical structure of Dracaena marginata 'bicolor' plants.
Starting from 20 th April till 20 th October, for both seasons, the plants grown in different soil media received monthly different treatments. A combination of NPK was added to soil in forms of (NPK,20:20:20) at the rate of 2 and 4 g pot -1 with foliar spray of either benzyladenine (BA) at the concentrations of (100, 150 ppm) or gibberellic acid (GA 3 ) at the concentrations of (150, 250 ppm) until dropping point. The control plants were sprayed only with tap water without fertilization.
On 20 th November for both seasons, the experiment was terminated and the vegetative growth traits were registered, including: plant height (cm), number of leaves/plant, stem diameter (mm), leaf area (cm 2 ), root length (cm), fresh and dry weights of leaves, stems and roots/plant. The chemical constituents were determined including: total chlorophylls (SPAD) in fresh leaf samples (Netto et al., 2005), total carbohydrates (%) in dried leaves and stems samples (Dubois et al., 1956). Another dried leaves and stems samples were digested for nutrients determination. The concentration of N (Horneck and Miller, 1998), P (Jackson, 1973), K (Horneck and Hanson, 1998), Ca and Mg (Estefan et al., 2013) and the contents of Cu, Fe, Mn and Zn (Chapman and Pratt 1961), beside B (Gupta, 1998) were determined.

Anatomical studies
At the end of vegetative growth, specimens of the leaves were taken and tissue specimens dissected to the size of the sample, and transferred to FAA solution at room temperature for 20 hours. After fixation, they were washed in 50% ethanol then dehydration was completed by transferring the sample to 100% butyl alcohol (changes, for one day each). Dehydrated sample were immersed into a 1:1 solution of 100% ethanol: xylene for 1 hour, then transferred into xylene for storage until infiltration with molten Paraplast in a 55-60 o C oven. The tissues were placed into wax 58 °C and placed into embedding molds in 7x7x5 mm disposable base molds (SLEE MAINS type MPS/P, Germany). The wax blocks containing the embedded tissue were sectioned using a rotary microtome (Leica RM 2125) with heavy duty high profile disposable microtome blades. (Ellis, 1976;Aiken et al., 1984). Images were acquired with a camera Leica ICC50 HD digital camera attached to a Leica motorized light microscope system. Averages of readings from 4 slides / treatment were calculated.
All previous data were subjected to analysis of variance (ANOVA). The effects of growing media (G), combined treatments of NPK with either BA or GA 3 concentrations (T), and their interactions was analyzed by two-ways randomized block ANOVA. The Least Significant Difference (L.S.D.) test at the 5% level was used to compare the means values of different parameters (Steel and Torrie, 1997).
V. 26, N o . 4, 2020 p. 545-561 Combined effeCts of nPK fertilizer with foliar aPPliCation of benzyladenine or GibberelliC aCid on Dracaena marginata 'biColor' Grown in different PottinG media 550 Table 4. Effect of growing media, combined NPK with foliar application of plant growth regulators treatments and the interactions between the two factors on fresh and dry weights of roots of Dracaena marginata during the 2018 and 2019 seasons. The favorable effect of peat-moss medium on enhancing vegetative growth may be attributed to its properties such as low pH and EC values in addition to its high water-holing capacity as indicated in (Table 1), this may be associated with water supply in sufficient quantities for turgidity and various positive metabolism including cells enlargement which in turn lead to stimulation of stem elongation (Fascella, 2015;Badran et al., 2017).

Effect of NPK with plant growth regulators treatments
Data in Tables (2-4) indicated that, the combined treatment of NPK with either BA or GA 3 concentrations resulted in significant increases in most of the vegetative growth parameters in terms of plant height, number of leaves/plant, leaf area, root length, fresh and dry weights of leaves, stems and roots as compared to the untreated plants.
In spite of this, it was notable that, the only parameter which insignificantly affected by integrated NPK rates with BA or GA 3 was stem diameter as compared to each other and control plants. Also, it was detected that, GA 3 was more effective than BA when they were combined with NPK at the same rates. These results are in agreement with the finding of the previous study on croton plants (Ibrahim et al., 2010) who reported that GA 3 treatments were more effective than BA on increasing plant height, leaf area, fresh and dry weights of stems, leaves and roots of croton plants.
Concerning the application NPK combined with the same concentrations of plant growth regulators (BA or GA 3 ), it was found that the lowest rate of NPK was more effective than the highest one; this may be due to plant sensitively against salt toxicity resulting from excessive fertilization rates. In both seasons, the highest values for most vegetative parameters were obtained from the plants treated with the combination of NPK at 2 g pot -1 with GA 3 at 250 ppm, while the lowest values were resulted from the control plants. Similar increases in the vegetative growth parameters due to application of NPK in combination with GA 3 had been authenticated by previous research (El-Sayed et al., 2016). Also, increases in growth parameters of Dendrobium densiflorum plants as a result of combined NPK with BA have been reported by prior study (Ngapui et al., 2018).
The above-mentioned results showed the important role of N, P and K as well as BA and GA 3 in the different physiological processes within the plant, which in turn affect the plant growth. Nitrogen is an important macroelement in organic molecules in plants including proteins, nucleic acids, purines, pyrimidines, co-enzymes (vitamins), and production of chlorophyll. Phosphorus is a component of sugar phosphates, nucleic acids, nucleotides, coenzymes, phospholipids, however potassium is essential in activation of enzymes and co-enzymes, proteins formation, photosynthesis, sugar transport, the uptake of other nutrients and their movement within the plant. The stimulated effect of BA may be due to its role in increasing cell division and enlargement, branches formation and overcoming the apical dominance leading to increase plant growth. The favorable effect of GA 3 on the plant growth may be attributed to its effect on stimulating the expression of enzymes involved in cell wall loosening and genes controlling cell division and also stimulating microtubule rearrangements associated with cell expansion (Satish and Manju, 2018). It was stated that the increase of plant growth by BA mainly associated with higher net assimilation rates and increased N content per unit leaf area (Di Benedetto and Galmarini, 2015), while increasing plant growth due to GA 3 may be attributed to its influence on increasing photosynthesis rate through increasing leaf surface (Lima et al., 2014). Moreover, increasing plant growth as a result of combined growth hormones with nutrient may be owing to physiological role of hormones and nutrient in synthesis of the plant phytochemicals through the action of various enzymes activity and protein synthesis (Tandel et al., 2018).

Effect of the interaction between growing media and NPK combined with plant growth regulators treatments
Data presented in Tables (2-4) revealed that generally, the plants grown in peat-moss or peat-moss + sand and received different NPK combined with BA and GA 3 treatments were significantly higher than those of the control plants. In both seasons, in most cases, the highest values recorded for most of the vegetative characteristics were obtained from plants grown in peat moss medium and received of 2 g NPK + GA 3 at 250 ppm as compared with the control which giving lowest values.

Chemical constituents
Total chlorophylls and total carbohydrates contents From the data Table (5) it's possibly discerned that, in both seasons, total chlorophyll in the leaves and total carbohydrates in the leaves and stems of Dracaena marginata ' Bicolor' were significantly higher in the plants grown in peat-moss+ sand medium than those grown in peat-moss alone. These findings are in accordance with the results obtained by previous studies which showed that a mixture peat-moss with sand increased chlorophyll content (EL-Quasni et al., 2014; Abd el Gayed and Attia, 2018). The data in the data Table (5) also showed that application of the combined NPK with either BA or GA 3 significantly increased total chlorophyll content in the leaves and total carbohydrates in leaves and stems as compared to the untreated plants. Similar results on increasing the contents of chlorophylls or carbohydrates due to application the combined NPK with GA 3 were reported by previous study (El-Sayed et al., 2016;Ghatas, 2016). Earlier studies (Tandel et al., 2018) reported increase in carbohydrates content due to combined urea with BA.
In both seasons, the lowest values of total chlorophyll content were obtained from plants grown in peat-moss medium without additional treatments, while the highest values were obtained from the plants grown in peat-moss + sand and received 2 g NPK + GA 3 at 250 ppm. In both seasons, the highest values of total carbohydrates were recorded in the plants grown in peat moss + sand and received 4 g NPK + GA 3 at 250 ppm, whereas the lowest values were determined in the plants grown in peat-moss + sand without treatments.

Macro elements
The chemical analysis of the leaves and stems (Tables 6 and 7) demonstrated that, generally, the uptake and accumulation of macro elements represented in N, P, K, Ca and Mg% were significantly higher in peat moss-grown plants than those grown in peat-moss + sand. However, in the first season there were no significant differences between the two tested growing media concerning the values of P and Mg% in stems and K% in the leaves.
The obtained results of increased N, P or K% in plants grown in peat moss alone compared to other media are similar to those found by prior research (Mousa et al., 2015). The obtained results of increased the accumulation of macro elements in peat moss-grown plants could be explained on the basis of high content of organic matter in peat moss maybe increase the activity of the beneficial microorganisms that play vital role in soil aeration and availability of the elements in rhizosphere to be absorbed by the plant roots. Moreover, its proper pH and EC values, high water-holing capacity may be had favorable effect in availability of nutrients and its accumulation in plant organs Pascual et al., 2018).
As for the effect of NPK in combination with BA and GA 3 the data in Tables (6 and 7) pointed out that, application of the combined treatments resulted in significant increases in N, P, K, Ca and Mg% in the leaves and stems compared to the control plants. However, in the first season plants treated with the combined treatments of 2 g NPK + BA at 100 or 150 ppm (T1and T2) had insignificantly higher values of P% in the stems than the control plants. The data also showed that, in most cases, the values of macro nutrients were increased steadily with increasing application rate of NPK and BA or GA 3 concentration compared the control plants. Furthermore, the combined NPK with GA 3 appeared to be more effective than combined with BA at the high concentration (250 ppm). These results of increased N, P or K% as a result of accompanied NPK with GA 3 treatments are in agreement with those obtained by prior research (El-Sayed et al., 2016;Ghatas, 2016), while increased N, P or K% due to the combined NPK with BA treatments are in harmony with prior study (Barman and Naik, 2017).
Regarding the interaction effect, data in Tables (6 and 7) showed that, plants grown in peat moss only or peat moss + sand and received different combined treatments of NPK with either BA or GA 3 resulted in significant increment of mineral nutrients percentage than those resulted from control plants. The data also revealed that, under both growing media GA 3 was more effective than BA when combined with NPK at the same rate.

Micro elements
The data in in Tables (8 and 9) showed that in most cases, the values of micro elements represented in Cu, Fe, Mn, Zn and B in the leaves and stems of plants grown in peat-moss + sand were higher than those of the plants grown in peatmoss alone. In the first season the accumulation of Zn and B in stems was the only exceptions which showed different trend, as peat moss grown plants produced significantly higher values than peat moss + sand medium.

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Combined effeCts of nPK fertilizer with foliar aPPliCation of benzyladenine or GibberelliC aCid on Dracaena marginata 'biColor' Grown in different PottinG media T1= Control, T2=2 g NPK + BA (100 ppm), T3=2 g NPK + BA (150 ppm), T4= 4 gNPK + BA (100 ppm), T5=4 g NPK + BA (150 ppm), T6=2 g NPK + GA 3 (150 ppm), T7=2 g NPK + GA 3 (250 ppm), T8=4 g NPK + GA 3 (150 ppm), T9= 4 g NPK + GA 3 (250 ppm) 2 G= Growing media, T= Treatments, G X T= Growing media X Treatments, different lower case letters in the columns and upper case letters in the rows for each variable indicate a significant difference at 5% level of significance by LSD test.    Combined effeCts of nPK fertilizer with foliar aPPliCation of benzyladenine or GibberelliC aCid on Dracaena marginata 'biColor' Grown in different PottinG media 558 In this respect previous finding  indicated that peat-grown plants were inferior to those grown in clay or rice straw in Fe, Cu and Mn contents in leaves of Gardenia jasminoides.
The data in in Tables (8 and 9) also cleared that the uptake and accumulation of Cu, Fe, Mn, Zn and B contents in the leaves and stems were markedly affected by the treatments of combined NPK with both BA and GA 3 as compared to the control plants. In both seasons, in most cases plants treated with any of the combined treatments had significantly higher values than the values obtained from untreated control plants. Also the data indicated that, GA 3 was more effective than BA when combined with NPK at the same rates.
Concerning the interaction effects, the data in Tables (8 and 9) revealed that plants received any treatments of combined NPK with either BA or GA 3 had significantly higher values of micro elements than those obtained from control plants. Overall, in both seasons the lowest values were obtained from plants grown in medium containing peat-moss only or peat-moss+ sand and received no treatments. While, the highest values were recorded with plants grown in medium of peat-moss+ sand and received combined treatments of 4 g NPK + GA 3 at 250 ppm.

Leaf anatomy
As shown in Fig.1 (a, b, and c) the leaves of Dracaena marginata are compound and composed of two little thicker upper and lower epidermis cells. They are covered with thick layer of cuticle particularly existing in the upper epidermis, meanwhile stomata were concentrated on the lower epidermis. Cross section in the leaves revealed that, the epidermis tissue cells (both upper and lower) of the control (Fig. a) were consisting of similar shape and size barrel, compactly arranged tiny cells and then become thicker and larger than control as shown in (Fig. b) as a result of the combined NPK with BA treatments. The application of NPK combined with GA 3 treatments resulted in the maximum size of the epidermis cells (Fig. c). Regarding the mesophyll tissue that consists of palisade and spongy parenchyma cells, in which the palisade lies just inner to the upper epidermis, and composed of elongated cells organized in two layers. The palisade cells region was compact and filled with chloroplasts which arranged along their radial walls, whereas the parenchyma cells tissue were present above and below the large vascular bundles. The spongy parenchyma cells region are loosely arranged, filled with chloroplasts and present below the palisade and extends up to the lower epidermis. It was remarkable that, the thickness of mesophyll tissue, which is specialized in photosynthetic and contains chloroplasts in palisade cells were visible and clear. The size of the spongy parenchyma tissue, air spaces of that treated with combined NPK with GA 3 became the largest as compared to both of combined NPK with BA treatments and control. Furthermore, vascular bundles of combined NPK with GA 3 treatment were larger and greater as shown in (Fig. c) as compared with the other treatments including control. These results are representing evidence of chlorophyll concentration, which was higher for the combined NPK with GA 3 treatment and that reflected the increased total carbohydrates percentage with the same previous treatment compared to control. This might be attributed to the increase in the chlorophylls synthesis and photosynthesis rate. The previous data are in conformity with the results obtained by (Ullah et al., 2017).

Conclusions
On the basis of earlier stated data, for the best vegetative growth and economic production of Dracaena marginata 'Bicolor', the plants advised to grow in a medium of peatmoss and supplied monthly with NPK fertilizer (20:20:20) at the rate of 2 g plant -1 along with foliar sprayed of GA 3 at the concentration 250 ppm.

Author Contribution
HAA: Responsible for implantation of the experiment according to established treatments, data collection, and data evaluation, arrangement of experimental data, statistical analysis, manuscript writing and corrections. ABE: Implantation of the experiment according to established treatments, data collection, statistical analysis, manuscript writing and corrections. MMA: Assistance in data collection, chemical analysis, statistical analysis, assistance in the preparation of manuscript.