Effect of eco-enzymes on vegetative and generative growth in several kipas putih soybean (Glycine max (L) Merr.) mutant lines

This research used soybeans resulting from gamma-ray radiation mutations of the Kipas Putih variety to obtain new superior varieties that are better than their parents. The research was carried out at the East Sector 2 Experimental Garden, as well as the Genetics and Plant Breeding Laboratory, Faculty of Agriculture, Syiah Kuala University from April to July 2023. This research used a Randomized Completely Block Design (RCBD) in a 2 x 4 factorial pattern with 3 replications so that there are 24 experimental units. The first factor is the concentration of ecoenzyme (K) which consists of 2 levels (without eco-enzyme and 10 ml L−1), while the second factor was the soybean line which consists of 4 levels, namely mutants B4, B7, B18, and the Kipas Putih (parent) variety. The results showed that the application of 10 ml L−1 eco-enzyme had an influence on the parameters of plant height, number of productive branches, number of pods, number of pithy pods, and percentage of pithy pods. The Kipas Putih G1 (B4) soybean mutant line was the best in terms of the number of pithy pods. The Kipas Putih G2 (B7) soybean mutant line was the best in the parameters of harvest age, flowering age, number of pods, and seed weight per plant. The best interaction of eco-enzyme with mutant lines was found in mutant lines G3 (B18) and G2 (B7) when given 10 ml L−1 of eco-enzyme based on the parameters of the number of productive branches and seed weight per plant.


Introduction
Soybeans originate from mainland China and have been cultivated since 2500 BC.Soybeans became known in Indonesia in the 16 th century [1].The increasing population of Indonesia every year means that the need for soybeans is also increasing, especially as a raw material for the food processing industry.The need for soybeans in Indonesia still depends on imports from various countries, because soybean production in Indonesia is not sufficient for people's needs.Therefore, various efforts to increase soybean production are very necessary, including new superior soybean varieties.
Kipas Putih is a local Acehnese soybean that was released as a National variety on November 3, 1992 based on a decision Minister of Agriculture: 619/Kpts/TP.240/11/92.This variety has a disadvantage because it has a small seed size, the weight of 100 seeds is only 12 g, so it is not popular in the tempe industry.The Kipas Putih variety has the potential to be developed into a new superior variety after genetic improvement through mutation techniques and is very suitable for existing land conditions in 1297 (2024) 012061 IOP Publishing doi:10.1088/1755-1315/1297/1/012061 2 Aceh [3].The results of previous research show that some of the best mutant lines have higher yield potential than their parent variety Kipas Putih.The B4 mutant line produces more than 2 tons ha -1 , and the B7 mutant line has a faster flowering age.The B18 line has the potential to become a new superior variety because it has a high yield potential, namely 3.02 tons ha -1 with a large seed size (4).
Agricultural systems that use excessive fertilizer can indeed double crop yields, but the negative impact can cause environmental damage in the form of decreasing soil fertility levels so that agricultural land becomes damaged [5].Based on this, it is necessary to make efforts to reduce the use of chemical fertilizers by providing environmentally friendly eco-enzymes.Eco-enzymes are liquid extracts produced from fermentation of vegetable and fruit residues mixed with sugar [6].[7] explained that the brown eco-enzyme liquid has a strong aroma.[8] wrote that the contents of eco-enzymes are acetic acid (H3COOH), nitrate (NO2), and carbon trioxide (CO3) which the soil needs as nutrients.[9] stated that acetic acid in eco-enzymes can damage organisms, so it can be used as an insecticide and pesticide.[10] explained that eco-enzyme can increase the production of edamame soybean plants, where the best treatment is in the ratio (1L eco-enzyme: 100 L water).It was further explained that eco-enzyme was given to edamame plants by watering the potted plants with 500 ml per plot during their growth period.[11] wrote that appkication of eco-enzymes up to a concentration of 10 ml L -1 also had a significant effect on the growth and production of shallots.
The effect of applying eco-enzymes has not been widely studied, especially on soybean plants [10].Based on the statement above, research was carried out regarding applying eco-enzymes to several Kipas Putih mutant soybean lines.This research aims to determine the effect of eco-enzymes on the vegetative and generative growth of several mutant lines of Kipas Putih soybeans.

Material
This research was carried out at the East Sector Experimental Garden of Syiah Kuala University from April to July 2023.Observations of the research results took placed at the Genetics and Plant Breeding Laboratory and the Greenhouse at the Faculty of Agriculture, Syiah Kuala University, Banda Aceh.The tools and materials used were polybags, hoes, rakes, raffia rope, 12 mesh soil sieve, hose, sprayer, measuring tape, scissors, tray, analytical scales, stakes 90 cm high, camera and stationery for making observations.The seeds used the Kipas Putih variety and mutant lines (B4, B7 and B18).Other materials used were cow manure, alluvial soil obtained from Indrapuri District, Aceh Besar Regency, basamid 98 gr, eco-enzymes (which made from banana peels, orange peels, grapefruit peels, watermelon peels, melon peels, dragon peels, and pineapple peels) with a ratio of 30 L of water, 9 kg of fruit peel and 3 kg of brown sugar, measuring cup, plastic clip, brown envelope, NPK fertilizer with a ratio of 16:16:16, furadan insecticide with the active ingredient Karbofuran 3%, Lannate 25 WP with active methomyl 25% and husk ash.

Method
This research used a Randomized Completely Block Design (RCBD) with a 2 x 4 factorial pattern with 3 replications.There were 8 treatment combinations, each combination was repeated 3 times, in total there were 24 experimental units.Each experimental unit consisted of 4 plants, so that in total there were 96 plants.For each experimental unit 3 plants were taken as samples.There were two factors studied, namely the concentration of ecoenzymes (K) and the soybean mutant line (G).The first factor, namely eco-enzyme concentration (K), consists of two levels (K0 = No ecoenzyme, K1 = 10 ml L -1 ), while the second factor were soybeans mutant line (G) which consists of 4 levels (G0 = Parent (Kipas Putih), G1= B4, G2= B7, G3= B18.).Kipas Putih soybean and mutant seeds line were obtained from the collection of Prof. Dr. Ir. Zuyasna, M.Sc.
Preparation of planting media began by preparing alluvial soil and cow manure in a ratio of 2:1, then sterilizing it using Basamid G with the active ingredient dazomet 98%.For every 1000 kg of soil, 100 grams of Basamid 98-G was given, then closed with plastic for 2 weeks in an airtight condition.The plastic cover was opened once a week with the aim of preventing the remaining methyl isothiocyanate gas from evaporating [12], then put into a polybag of 35 x 40 cm and arranged in the field.
Eco-enzymes were obtained from the USK PPLH-SDA collection (Syiah Kuala University Environmental and Natural Resources Research Center).Eco-enzyme was made by preparing 9 kg of fresh fruit peel that was not rotten (banana peel, orange peel, grapefruit peel, watermelon peel, melon peel, dragon fruit peel, pineapple peel, and mango peel), 3 kg brown sugar, and 30 liters of water.The fresh fruit peel and brown sugar were placed in a container filled with water and stirred until evenly mixed.The container was then tightly closed and stored for 3 months starting from January 3 to April 3, 2023.During storage, the eco-enzyme liquid was opened every week and stirred to release fermentation gas.After 3 months of storage, the eco-enzyme liquid was ready for use.
Soybeans were planted with a distance between polybags of 40 cm x 30 cm.Soybean seeds were planted one by one to a depth of 2 cm.Each planting hole was filled with 2 hollowed-out seeds.The application of eco-enzyme was carried out once a week for five weeks with a concentration of 10 ml of eco-enzyme dissolved in water until the volume becomes 1000 ml, then given to the plants as much as 200 ml per polybag (concentration 10 ml L -1 ).The chemical fertilizer given was NPK fertilizer (16:16:16) as basic fertilizer, namely at a dose of 2 g L -1 of water, application was carried out two weeks after planting.Pest and disease control was carried out mechanically and chemically every week.
Weeding was done intensively until the plant entered the generative phase.Chemical pest control was carried out if symptoms of the disease were visible on the plant.If the plant showed symptoms of caterpillar or insect attack, it was given the insecticide Lannate 25 WP with the active ingredient 25% methomyl in a concentration of 2 g L -1 of water.Parameter observations were made on plant height at 2, 4, 6, 8, and 10 weeks after planting (WAP), number of productive branches, flowering age, harvest age, total number of pods, number of pithy pods, seed weight per plant, and percentage of pithy pods.

Application of ecoenzymes to several Kipas Putih soybean mutants and their effects on vegetative and generative growth 3.1.1. Plant height and number of productive branches
The best plant height parameters at the ages of 2, 4, 6, and 10 WAP were found in the G3 (B18) mutant line with 10 ml L -1 of eco-enzyme as presented in Figure 1.Statistically, eco-enzyme treatment had a significant effect on plant height parameters and the number of productive branches.The highest value was found when applying 10 ml L -1 of eco-enzyme, because the N content in the soil was sufficient.The response to the addition of N elements through eco-enzyme fertilization could be seen, as shows in Table 1.
Figure 1.The plant height in several mutant lines: A. without applying eco-enzim; B. addition of ecoenzim Note: Numbers followed by the same letter in the same column are not significantly different at the 5% level (DNMRT 0.05) (13), stated that one of the factors controlling plant growth is nitrogen (N), which functions in protein formation and increases plant vegetative growth such as plant height and number of productive branches.(14), stated that plants use more elements (N) which function for shoot growth than root growth, so that this affects the growth of plant height and the number of productive branches.The N content in ecoenzyme is very low, namely 0.07%, but the greater the concentration of eco-enzyme given to the plants, the more N is added so that it can be absorbed and cause plants to grow taller.Statistically, genotype does not affect plant height and number of productive branches.According to [15] this is influenced by genetic factors.High diversity in the generative phase shows that character is more influenced by genetic factors.Variations in plant height occur because each genotype has different genetic factors and characteristics.Apart from that, the differences in genotypic characters that appear in each genotype are caused by differences in the genes that regulate these characters.Each gene has its task to grow and regulate various types of characters in an organism.

Flowering time and harvest time
Table 2 shows that the fastest plants flowering were in the 10 ml L -1 treatment of eco-enzym (40.67 DAP), which was not statistically different from without the application of eco-enzyme.The fastest average age of harvest was found in the 10 ml L -1 treatment (91.86 DAP), which was statistically not different from without the application of eco-enzymes.The fastest plants to flower tended to be found in the G2 (B7) and G3 (B18) mutant lines, namely at 40.17 DAP, although statistically not different from the other mutant line.The fastest plants to harvest tended to be those treated with the G2 (B7) mutant line (91.83DAP), although statistically not different from the other mutant line.The number of productive branches tended to be higher in the treatment of the G3 (B18) mutant line (7.00 branches), although statistically not different from other mutant lines.
The parameters of flowering age and harvest age are not significantly different, this is because flowering age and harvest age are more influenced by genotype.[16], stated that the harvest time for each plant genotype depends on the flowering age and also the genetic factors of each genotype.The genotypes tested had different responses to genetic factors and genotype responses in response to environmental conditions on the planted land, resulting in differences in the appearance of flowers for each genotype.Further more [17], wrote that each plant genotype has different growth characteristics due to differences in the plant's genetic characteristics, so that the longer vegetative growth takes, the longer the time for flowers to appear.3 shows that the total number of pods, the number of pithy pods, and the percentage of pithy pods were highest in the treatment with eco-enzymes (10 ml L -1 ), namely respectively 92.72, 81.58, 88.16, 14.48 g which was significantly different from without eco-enzyme.The highest seeds weight per plant was found in the treatment with eco-enzymes (10 ml L -1 ), namely 14.48 g, although this was not significantly different from the treatment without eco-enzymes.The best total pod number parameter was found in the G2 (B7) mutant line with a value of 91.17 piece pods.The parameter for the number of pithy pods tended to be better in the G1 (B4) mutant line, namely 75.33 piece pods, which was not statistically significant different from the other mutant line treatments.The percentage of pithy pods tended to be better in the G3 (B18) mutant line, namely 89.50 piece pods, although statistically it was not significant different from the other mutant line treatments.The highest average seeds weight per plant was found in the G2 (B7) mutant line, namely 14.74 g, which was significantly different from the G0 (parent).The provision of eco-enzymes had a significant effect on the total number of pods, the number of pithy pods, and the percentage of pithy pods, this was because the nutrients N and K contained in the 1297 (2024) 012061 IOP Publishing doi:10.1088/1755-1315/1297/1/0120616 eco-enzyme used in the fruit skin had increased significantly.Physiologists believe that increasing the number of pods and seeds in plants can occur through the mechanism of carbohydrate metabolism from photosynthesis.Increasing the amount of Ca and Mg from application of this eco-enzyme can increase the number and weight of soybean pods.By increasing the amount of Mg, one of the elements that make up chlorophyll, which can help the photosynthesis process become faster with the Adenosine triphosphate (ATP) reaction, so that the number of Kipas Putih soybean pods will increase.Pod formation in Kipas Putih soybean plants is closely related to the process of plant photosynthesis, the formation of proteins, enzymes, hormones and carbohydrates to encourage cell enlargement and elongation so that plants will reproduce and experience optimal production, this can happen with the help of eco-enzyme catalysts [10].
Statistically, the total number of pods and seeds weight per plant have a significant effect on the treatment of mutant soybean lines because seed size was influenced by plant genetic factors.[18], wrote that seed size character is one of the traits that is influenced by genetic factors and the plant's environment during seed development.

The interaction of mutant lines and eco-enzymes on the vegetative and generative growth of soybean plants (Glycine max (L). Merr) 3.2.1. Number of productive branches
Table 4 shows that treatment without eco-enzyme on the number of productive branches was statistically the best in the G2 (B7) mutant line with a value of 6.89 branches.The best eco-enzyme treatment of 10 ml L -1 was found in the G3 (B18) mutant strain with a value of 7.56 branches.Statistically, the number of productive branches had a very significant effect on the treatment of mutant lines by addition of 10 ml L -1 of eco-enzyme to the G3 (B18) mutant line, because the N content in the soil was very sufficient, so the response to the addition of N elements through eco-enzyme was very visible but there was differences in response patterns between genotypes to whether eco-enzymes were given or not given eco-enzymes.This effect can be explained because genotypes have different abilities in absorbing nutrients in the soil.[19], wrote that genotypes have different abilities to adapt to the plant's environment 3.2.2.Seeds weight per plant (g) Table 5 shows that treatment without giving eco-enzymes on the weight of planted seeds was statistically the best in the G3 (B18) mutant line with a value of 15.01 g.The best 10 ml L -1 ecoenzyme treatment was found in the G2 (B7) mutant strain with a value of 15.70 g.
Seeds weight parameters were very significantly different due to the interaction of eco-enzyme application and Kipas Putih soybean mutant lines.The application of eco-enzymes did not have a big influence in determining seed weight per plant but was a result of genetic factors.[18], wrote that seed size is one of the traits that is influenced by genetic factors and the plant's environment during seed development.

Conclusions
Giving 10 ml L -1 of eco-enzyme produced the best parameters for plant height, number of productive branches, the total number of pods, number of pithy pods, and percentage of the pithy pods.The Kipas Putih G1 (B4) soybean mutant line was the best in terms of the number of pithy pods.The Kipas Putih soybean mutant line G2 (B7) was the best in terms of the parameters of harvest age, flowering age, the total number of pods, and seed weight per plant.The Kipas Putih soybean mutant line G3 (B18) was the best in terms of plant height, number of productive branches, flowering time, and percentage of pithy pods.The best interaction of eco-enzyme with mutant lines was found in mutant lines G3 (B18) and G2 (B7) when applying 10 ml L -1 of eco-enzyme based on the parameters of the number of productive branches and seed weight per plant.

Table 1 .
Average plant height at 2, 4, 6, 8, and 10 WAP and the number of productive branches of the Kipas Putih soybean mutant line when applied with eco-enzymes

Table 2 .
Average flowering age and harvest age of Kipas Putih based on genotype and aplication of eco-enzyme Total number of pods, number of pithy pods, percentage of pithy pods and seed weight per plant Table

Table 3 .
Average the total number of pods, number of pithy pods, percentage of pithy pods and seeds weight per plant of Kipas Putih Soybean plants on aplication of ecoenzyme

Table 4 .
The average number of productive of soybeans due to the interaction of providing eco-enzymes and Kipas Putih soybean mutant lines

Table 5 .
Average of seeds weigh per plant due to the interaction of providing eco-enzymes and Kipas Putih soybean mutant lines Numbers followed by the same letter (capital letters read vertically and lower case letters read horizontally) are not significantly different at the 5% level (DNMRT 0.05)