Prediction of leaf N in centipedegrass by a chlorophyll meter

The persistence of color response to fertilizer treatments is an important criterion of turfgrass performance. Centipedegrass (Eremochloa ophiuroides (Munro) Hack.) is widely used throughout subtropical region, and was selectedas the subject of this study. Four N treatments were applied, with NH4NO3monthly atan application rate of 0, 2.5(1.4), 5.0(2.8), and 10.0(5.6) g-N/m 2 (oz). A slow-release fertilizer applied as Osmocote at an application rate of 7.5g-N/m 2 was added to each treatment. The trial was conducted in the greenhouse for 12 weeks. Chlorophyll content, N content, leaf area, plant biomass measurement, and growth characteristics were used to determine turfgrass growth qualities. From the results we found that highest average plant height, leaf length, leaf width, shoot dry weight (DW), and root DW were obtained with all fertilizer products application than without N fertilizer input. Regardless ofN fertilizer concentrations, the first leaf, and third leaf CMR increased with N treatments compared to the control in this study. Centipedegrass CMR closely corresponded to N application concentration. The experimental results revealed that average sufficiency index (142) showed an adequate amount of tissue N supplied at this stage of centipedegrass growth. Linear regression of leaf chlorophyll content and CMR values was found in this study (r 2 =0.9436, P<0.01). This research suggested that the chlorophyll meter could be useful to directly understand the tissue N in centipedegrass. The linear regression of dry weight-based or area-based N concentration on leaf CMR values was highly significant (P<0.01). The results also illustrated that CMR values correlated with Ndw (r 2 =0.9034) (N content/dry weight) better than with Na (r 2 =0.7611) (N content/leaf area), indicating that CMR estimated Ndw better than Na. Thus, this investigation suggested that Ndw is the major contributor to variation in SPDA-502 chlorophyll meter readings. Chlorophyll meter measurement can offer an alternative to the tissue test, and can aid in determining fertilizer N recommendations for centipedegrass was demonstrated in this study.


INTRODUCTION
Centipedegrass is a native turfgrass of Taiwan and is one of the most popular turfgrass species widely used in other subtropical regions (Turgeon, 1991).Efficient use of N fertilizer is important to economical and environmental turfgrass production, and to ground and surface water quality (Emmons, 1995;Stevenson, 1986).Assessment of fertilizer product performance under controlled conditions is a potentially useful method for prescreening turfgrass response to fertilizer treatment before conducting actual field trials (Horst et al., 1994;Miller and Thomas, 2003;Kuo et al., 2005).While the field performance of various N sources is difficult to forecast future performance given unknown climate (Takebe, 1990).In field, the turfgrass quality after treated with N fertilizer is still determined largely through visual assessment.
However, visual assessment is generally imprecise, not definitive, and dependent on evaluator prejudice (Ichie et al., 2002).Turfgrass quality cannot be measured in the same way as that of other agricultural crops (Morris, 2001).More efficient, low-cost, and non-destructive methods of pre-screening turfgrass growth are required to help tackle the problem of excessive fertilizer use in intensively cultivated areas (Bell et al., 2004).However, the traditional method of extraction and quantifying plant N content is both times consuming and destructive.Recently, reflectance measurements are popular using in prediction fertilizer status in crops (Trenholm and Unruh, 2005).In addition, more promising method of chlorophyll meters enable quick and easy measurement of leaf blade greenness, which is an indicator of shoot N and chlorophyll content (Rodriguez et al., 2000;Takebe et al., 1990;Turner and Jund, 1991;Wood et al., 1992a).Chlorophyll meters also display promise as a tool for improving N management (Peterson et al., 1996;Stevens and Hefner, 1999).Numerous researchers have proved the positively linear relationship between tissue N status and chlorophyll meter readings of some crops (Fox et al., 1994;Kantety et al., 1996;Minotti et al., 1994;Wood et al., 1992b).
Moreover, conditions for the use of chlorophyll meters (Minolta SPAD-502) recently have been clarified for wheat (Denuit et al., 2002), woody plants (Ichie et al., 2002), rice (Ramesh et al., 2002), peanut (Nageswara Rao et al., 2001), and barley (Giunta et al., 2002).However, the application of chlorophyll meters to more economically valuable and popular warm-season species, such as centipedegrass has not been explored.Turfgrass response to different fertilizer products has been evaluated in a numbers of different species but hardly found in centipedegrass.The present experiment comprised two parts.First, centipedegrass response to fertilizer rates and sources was screened; second, the relationship between Minolta SPAD-502 chlorophyll meter readings and plant biomass, plant growth characteristics, leaf area, and N status in leaf was investigated.

MATERIALS AND METHODS
Seeds of centipedegrass (from the Rose Extension Center, Taipei, Taiwan) were grown in a 15 cm diameter X 20 cm deep plastic pot containing loam, peat moss and vermiculite (2:2:1 The greenhouse study was conducted in the environmental controlled greenhouse (22/20°C day/night, 3.64MJ m -2 d -1 ) (Model LX-102 potable light meter, Alfa Electronics inc., NJ.) located in the campus of Taiwan University, Taipei.Pots were watered twice weekly.Different sources and rates of N were applied every 4 weeks as Ammonium Nitrate (NH4NO3; Tai-Fertilizer Company, Taipei, Taiwan) at an application rate of 0 , 2.5(1.4), 5.0(2.8), and  10.0(5.6)g-N/m 2 (oz) after seeds germination.A slow-release fertilizer applied as Osmocote (14N: 6.2P: 11.6K; Tai-Ho Company, Taipei, Taiwan), a resin-coated fertilizer, at an application rate of 7.5(4.2) g-N/m 2 (oz) was included in each treatment for once only at the first time treatment.
Additionally, a modified N free full-strength Johnson's nutrient solution was applied weekly to prevent deficiency of the other essential elements (Table 1).Potable water was used as a control.The first and third fully expanded leaves (those with collars surrounding the stem) from the apex of the plant were randomly sampled from each pot for CMR measurement (Model Minolta SPAD-502, Minolta Co. Ltd., Japan).Twenty-five randomly selected samples from each portion of leaves were measured, thus there were totally 50 samples recorded for CMR and chlorophyll content measurement (Mode U-01 UV/Visible Spectrophotometer, Hitachi Ltd. Japan)to study the correlation between them.The N sufficiency index (defined as the SPAD value of a plant receiving fertilizer divided by the SPAD value of a plant not receiving fertilizer times 100) was calculated.The CMR measurement was based on the difference between light attenuation at 430 nm and 750 nm, with no transmittance.Moreover, the length and width of the third fully expanded leaf was measured before harvested.The SLW of the third fully expanded leaf was calculated as the ratio of leaf weight to area.Leaf area was measured using LI-COR area meter, (Model Li-3000, Alfa Electronics inc., New Jersey, USA).Leaf N content was analyzed by semi-micro Kjeldahl digestion and distillation (Kuo et al., 1999).Leaf N content was expressed based on both the dry weight (Ndw) (N content/dry weight) and leaf area (Na) (N content/leaf area) to compare which one is more promising as indicator for presenting leaf N status when compare to CMR.Harvested sample seedlings of centipedegrass were divided into root and shoot components and oven dried at 70°C for 48 h after CMR measured.The root and shoot dry weight (DW) was recorded.The experiments were repeated once lasting for 12 weeks.The sample pots of both experiments in the greenhouse bench were completely randomized designed.Mean separation was evaluated at the 0.05 probability level using Duncan's multiple range tests (Statistix 8, Analytical Software, Orlando, Florida, USA).Since the result of two experiments were similar, so only one experimental result was shown in this study.

RESULTS AND DISCUSSION
Highest average plant height, leaf length, leaf width, shoot DW, and root DW were obtained with all fertilizer products application than without N fertilizer input (Table 2).Treatments receiving the higher rate of N only had significantly higher plant height but not significantly responded to their leaf length, leaf width, shoot DW, and root DW than slow-release N treatment only, in spite of the higher rate of fast-release fertilizer applications.These results revealed that fast-release fertilizer treatment is shorter-term performance and do not contributed to root biomass of centipedegrass.Slowrelease fertilizer alone provides longer-term release of N, less potential for leaching if misapplied, and less danger of turfgrass burn than do fast-release fertilizer sources.Regardless of N fertilizer concentrations, the first leaf, and third leaf of CMR increased with N treatments compared to the control in this study (Table 3).Centipedegrass CMR closely corresponded to N application concentration (Table 3).The relationship between adequately fertilized turf and N sufficiency index must be established in turfgrass research (Idso et al., 1996).Our experiment revealed that average sufficiency index (142) showed an adequate amount of tissue N supplied at this stage of centipedegrass growth (Table 3).
However, a high level of N application did not significantly influence CMR values.The analysis of N content of different rate of fertilizer treatments found that significantly influenced both dry weight and leaf area  bases (Table 3).Figure 1 illustrates the linear regression of leaf chlorophyll content and CMR values (r 2 =0.9436,P<0.01).From Figure 1, chlorophyll content was much responded to CMR and this finding agreed with that of Tunner and Jund (1991).Due to this relatively consistent degree of agreement between CMR values and plant chlorophyll content, it appears that the Minolta SPAD chlorophyll meter can be useful to directly understand the tissue N in centipedegrass.The linear regression of dry weight-based or area-based N concentration on leaf CMR values was highly significant (P<0.01).better than with N a (r 2 =0.7611), indicating that CMR estimated N dw better than N a .Thus, this investigation suggested that N dw is the major contributor to variation in SPDA-502 chlorophyll meter readings.Thus, the CMR can accurately describe the N status of centipedegrass in the field, and thus determine whether N fertilizer application is required.Idso et al. (1996) also found a close relationship between leaf chlorophyll a, b, carotenoids, and xanthophylls and SPAD-502 chlorophyll meter readings in sour orange tree leaves.This relationship is confirmed by the fact that N is a key element in chlorophyll molecules, and N contributes to plant shoot dry weight.Chlorophyll meter measurement can offer an alternative to the tissue test, and can aid in determining fertilizer N recommendations for turfgrass.The present work demonstrated that using a chlorophyll meter to measure centipedegrass growth and development is both simple and effective.Future field experiments will determine various application intervals and different application rates for use in greenhouse studies of centipedegrass.

Figure 1 .
Figure 1.Linear regression between leaf chlorophyll content and chlorophyll meter readings in centipedegrass.

Figure 2 .
Figure 2. Linear regression of dry weight-based (Ndw) and leaf area-based (Na) leaf N content on chlorophyll meter readings in centipedegrass.

Table 1 .
Composition of modified N-free Johnson's solution.

Table 2 .
Effect of N applications on plant height, leaf length, leaf width, shoot DWand root DW of centipedegrass.
ab*Values in a column followed by the same letter are not significantly different at 5% level, by Duncan's multiple range tests.

Table 3 .
Effect of N applications on average CMR value of first and third fully expanded leaf, sufficiency index, dry weight based N (Ndw), and leaf area based N (Na) of centipedegrass.