Bulletin of The Iwate Agricultural Research Center No.12

Effect of Nitrogen Uptake by Spinach (Spinacia oleracea L.) under Application of Manure Compost and Evaluation Method of Available Nitrogen for this crop in the Soils
@In Iwate Prefecture, there is an urgent need for proper soil management that effectively utilizes the large amount of organic waste produced from livestock. On the other hand, spinach is cultivated throughout Iwate Prefecture. I applied compost made from livestock waste into spinach production fields. The research consisted of the following four objectives:
@i) To observe the growth and nitrogen (N) absorption by spinach grown in fields applied with this compost and as well as to monitor any NO3 leaching from the surface soil to ground water in these fields.
@ii) To understand the mechanism in which spinach absorbs and uses available N in soil.
@iii) To propose a simple evaluation method of available N in soil that is suitable to the cultivation of spinach.
@iv) To propose a suggestion of an appropriate nitrogen level for soil treated with compost
@In a 4-year continuous experiment, the effects of a sawdust-like compost made from cattle feces on spinach growth were evaluated in a field covered by a plastic top to prevent rainfall. Spinach was harvested 2-4 times per year to analyze its growth and nitrogen absorption. The chemical fertilizer zone was fertilized annually with 16-20g/m2 of ammonium nitrate, 20-24g/m2 of monocalcium phosphate, and 16-20 g/m2 of potassium chloride. The compost zone was fertilized annually with 45gN/m2 of compost made of cattle feces 2 weeks before the first sowing of seeds, and was not fertilized after the 2nd sowing.
@The results are as follows:
@1) Plant length, leaf width, and number of leaves of spinach grown in the compost fertilizer zone were greater than those of spinach grown in the chemical fertilizer zone.
@2) Dry matter yield and nitrogen absorption was twice as high in the compost fertilizer zone when compared to the chemical fertilizer zone.
@3) Inorganic nitrogen content in soil that had spinach planted in it was lower in the compost zone than the chemical fertilizer zone. This tendency was the same through the entire 4-year experiment, and the density of inorganic nitrogen content in compost-added soil shifted to a level lower than the chemical fertilizer-added soil.
@4) Moreover, the content of NO3 within the spinach was found to be much lower in the compost zone than the chemical fertilizer zone. In other words, the content of NO3 increased greatly in soil treated with chemical fertilizer, so the content of NO3 within the spinach was also found to be high.
@5) Less inorganic nitrogen and all other nitrogen content leached below ground in the compost zone than the chemical fertilizer zone.
@These results show that spinach growth and nitrogen absorption are greater when compost is used as fertilizer, even though less inorganic nitrogen is found in the soil when compared to chemical fertilizer. In other words, the spinach growth is not responding to inorganic nitrogen in the soil.
@It is clear that spinach responds to soil fertilized with organic compost, but is thought that the most influential organic nitrogen absorbed by spinach is PEON, a protein-like organic nitrogen that is extracted from soil with a 1/15M phosphate buffer (pH 7.0). PEON has a molecular weight of 8000, can absorb UV, and acts similarly to protein (by the Bradford method of protein assay). We extracted PEON from the soil and injected it into rabbits, made PEON antibodies, and confirmed that it was possible to quantify PEON in soil. We took xylem sap from the spinach and used anti-PEON antibodies to quantify PEON content using the enzyme-linked immune sorbent assay (ELISA) method. The xylem sap of spinach fertilized with compost showed a high response to the anti-PEON antibodies, while the xylem sap of spinach fertilized with chemical fertilizer did not show much of a response at all. This result shows that spinach has a great ability to directly absorb PEON. Moreover, this explains the growth and nitrogen absorption in spinach grown in soil fertilized with compost, as we understand that this is through not just inorganic nitrogen but PEON as well.
@PEON has been shown to play a big part in the growth of spinach, but other organic nitrogen must be considered. We developed a sequential extraction method (A, B) to understand the form of nitrogen in soil that can be used by spinach.
@1) Sequential Extraction Method A: Water, 10% potassium chloride solution, 1M acetic acid, 1/15M phosphate buffer solution, 0.4M sulfuric acid, and 1M sodium hydroxide solution in sequence
@2) Sequential Extraction Method B: Water, 10% potassium chloride solution, 0.01M acetic acid, 0.1M sulfuric acid, 0.2M sulfuric acid, 0.4M sulfuric acid, and 0.5M sulfuric acid in sequence
@NO3 was found during the water extraction, and ammonia nitrate in the potassium chloride solution. Next, organic nitrogen was found in sequential extraction method A with acetic acid, phosphate buffer solution, and sulfuric acid, but according to HP-SEC, the holding time levels of this organic nitrogen peaked just like PEON.
@The organic nitrogen extracted through the various amounts of sulfuric acid content in method B all had a comparatively average molecular weight, which was the same as PEON at MW=8,000. Also, it was clear that this organic nitrogen was bonded to iron and aluminum. In other words, it was shown that nitrogen from compost-treated soil is a multilayered construction aided by iron and aluminum as PEON. Citric acid secreted from the roots of plants and oxalic acid secreted from the spinach are organic acids that dissolve PEON, which the spinach directly absorb.
@Because the organic nitrogen extracted from the soil through the sequential extraction methods is PEON or PEON-like material, we used sequential extraction method A to evaluate the organic nitrogen level in soil used to grow plants. We cultivated spinach and corn in greenhouse soil without fertilizer. We took xylem sap from each plant harvest, and analyzed the molecular weight allocation using HP-SEC. A peak in retention time in the xylem sap of spinach cultivated in soil was found that was similar to the organic nitrogen extracted with the 0.4M sulfuric acid. Hydroponically-grown spinach fed with inorganic nutrients did not have this peak. Moreover, we analyzed the formulation of organic nitrogen used by spinach using sequential extraction methods. We used the same sequential extraction method on soil that had been used to grow corn. The results showed that the fraction of nitrogen that had been extracted with 0.4M sulfuric acid from spinach soil had decreased. In other words, the spinach was shown to have used the nitrogen even in the fraction extracted from 0.4M sulfuric acid.
@As an estimation method of possible nitrogen content in soil used for the cultivation of spinach, we used the 0.4M sulfuric acid extraction method to verify the relationship between light absorption and extracted nitrogen content in 280nm of extracted liquid. There we found an extremely high level of correlation between the two. Following these results, we calculated the estimated organic nitrogen content from the light absorption level (280nm) within the 0.4M sulfuric acid extraction solution, and deliberated the relationship to the nitrogen absorption content in spinach that had been grown without fertilizer (3 harvests a year). As a result, it was shown that there is a tendency for the nitrogen absorption content in spinach to increase when the estimated organic nitrogen content in soil increases, but it was also found that nitrogen absorption efficiency in spinach decreases when estimated organic nitrogen content in the farm field is over 540mg/kg. We used chemical fertilizers or compost to cultivate spinach annually in fields with different estimated nitrogen contents, and the compost zone yielded more spinach and reduced the amount of nitrate. On the other hand, even though the amount of nitrate within the compost spinach decreased when compared to spinach grown with chemical fertilizer, nitrate levels rose above 3,000mg/kg when fields had more than 540mg/kg of estimated organic nitrogen content. We propose that a simple method to determine the amount of chemical fertilizer or compost to be used can be efficiently determined measuring sulfate-extractable organic nitrogen content in soil with 280nm of light absorption.
Breeding New Varieties@of Japanese Barnyard Millet@with Shorter Culms and Low Amylose Content: "Nebarikko 1", "Nebarikko 2" and "Nebarikko 3"
Shinsuke NAKAJO, Satoshi HASEGAWA, Hiroshi YOSHIDA, Shoji URUSHIBARA, Akira ABE, Tomoko ABE, Nobuhisa FUKUNISHI, Hiromichi RYUTOU and Yasumi OHSHIMIZU
@Three new types of Japanese barnyard millet (Echinochloa esculenta ((A.Braun)) H.Scholz ) were developed by applying gamma-ray and heavy-ion beam irradiation to a low amylase content landrace called eMojappe,f which is collected from Iwaizumi-cho, a town located in the mountainous area of Iwate Prefecture. 'Nebarikko 1' and 'Nebarikko 3' were derived from 600 Gy of gamma-ray irradiation, and 'Nebarikko2' was derived from 20 Gy of heavy-ion beam irradiation. All of these varieties have shorter culms than the parent 'Mojappe' but similar amylase content levels. However, they have different maturity rates and other morphological traits.
@'Nebarikko 1' matures earlier than 'Mojappe' with a shorter awn. It is presumed that the yield of 'Nebarikko 1' will be the same as that of the original variety. However, the culm length of this variety is influenced by environmental conditions, thus this variety is mainly recommended as an upland farming variety of barnyard millet in the northern part of Iwate.
@'Nebarikko 2' belongs to middle-maturity group, identical to 'Mojappe'. Although this variety has a small panicle, it has a large number of them, which results in a yielding level comparable to 'Mojappe'. Additionally, we expect that its awn-less kernel is suitable for mechanical hulling and sawing. The length of its culms also have the lowest rate of fluctuation within the three cultivars.
@'Nebarikko 3' is a late-maturing variety, and has broad, deep green leaf blades. The yield of this variety is lower than 'Mojappe'. On the other hand, its culm is thicker and more rigid. 'Nebarikko 2' and 'Nebarikko 3' are expected to be suitable for machine-planting and harvesting in paddy fields. Considering their maturity rates and intrinsic qualities, 'Nebarikko 1' is suitable for upland fields in the northern area of the prefecture, 'Nebarikko 2' can be cultivated all over the prefecture, and 'Nebarikko 3' can be cultivated in the central and southern areas of Iwate. As such, there are plans to increase its cultivation as a paddy field-based cultivar within Iwate.
Development of Methods that Continuously Induce Flowering in Strawberry Nursery Plants by Applying Nitrogen Fertilizer with a Short-day Night-chilling Treatmen‚”
Takuya FUJIO, Yuji SASAKI and Hiroshi SATO
@When forcing strawberries in a strawberry forcing culture, increasing yearly fruit yield is a challenge because there is a harvest delay between the 1st and 2nd flower truss. Therefore, using the forced cultivar eSachinokaf nursery plants, cultivation methods and other growing methods were examined that grow flower buds by inducing the 2nd flower truss.
@1. Induction of the 2nd truss was caused by extending the period of short-day treatment to more than 60 days, and flowers began to bud. However, using only the short-day method was an unstable method of efficiently inducting flowers, and induction of the 2nd truss was determined to need a short-day night chilling method treatment.
@2. During the short-day night chilling method, we applied nitrogen fertilizer after the 1st truss flower buds grew, which encouraged the induction and formation of the 2nd truss. The nitrogen content in additional liquid fertilizer was 50-75mgN/L, and was applied once a day at about 100mL per plant. This resulted in a higher ratio of effective plants. Additionally, the capacity in the soil was determined to be 360mL per plant, and soil with less capacity will be unsuitable when using the same methods.
3. Plants that had not yet had their 2nd truss appear were also mixed in during the short-day night chilling method, but when planted in late August, the effects high temperatures and long-day conditions were lessened, and flower bud induction could be achieved. But three or more days of high temperature and long-day conditions even within the short-day night chilling treatment will strongly inhibit 2nd truss induction.
4. In areas similar to the cool summer conditions of test site, such as the coastal area of Iwate Prefecture, night temperatures during the night-chilling treatment are good at 22 deg C, and the method will result in a success rate of over 80%. Additionally, if night temperatures become even cooler, flower buds will be induced sooner, but the cooling load will increase.
5. During cultivation, thinning out fruits increased average fruit weight. However, the fruit yield was not affected by fruit thinning, electric lighting periods, and concentration of culture solutions.
6. After planting nursery plants in late August that have had 62 days of the short-day and night chilling method and additional fertilizer applied, these plants will possibly have a higher yield than forcing cultures planted at the same time (during harvest in December). Additionally, because there is continuous growth between the 1st and 2nd flower truss, yield is increased at harvest time because there is no need for rest in between.
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