HPIC analysis of
phosphite
treated turfgrass
John Dempsey BSc (Hons)
HPIC analysis of phosphite treated turfgrass
Introduction
Potassium phosphite is regularly applied to amenity turfgrasses as part of routine
maintenance programs. Phosphite (PO33-) is derived from the alkali metal salts of
phosphorous acid - H3PO3, which when in contact with water forms phosphonic acid(Guest
and Grant, 1991; Rickard, 2000). The pH of this acid needs to be modified prior to use to
prevent phytotoxicity (Ouimette and Coffey, 1988) and this is most commonly done with the
addition of potassium hydroxide (KOH), forming potassium dihydrogen phosphite (KH2PO3)
or dipotassium hydrogen phosphite (K2HPO3). These compounds are the active substances in
numerous phosphite products which have been used in commercial horticulture and turfgrass
management since the 1980’s (Landschoot and Cook, 2005). Initially phosphite was used to
control Oomycete pathogens –Pythium spp.(Cook et al., 2006; Schroetter et al., 2006). It was
later found that combined with Mancozeb, a dithiocarbamate fungicide (Beard and Oshikazu,
1997) phosphite could improve significantly turf quality and also control Summer Decline of
bentgrass (Cook et al., 2006). At present phosphite products are extensively marketed and
used in amenity turfgrass management as both fungicides and biostimulants and while
research into the efficacy of phosphite in these areas has been carried out (Sanders, 1983;
Vincelli and Dixon, 2005; Horvath et al., 2006; Cook et al., 2009), there is none specifically
into phosphites assimilation and translocation in turfgrass.
This research was carried out to determine the assimilation rate, translocation and
accumulation of phosphite in a commonly used cool-season turfgrass –Agrostis stolonifera.
Turfgrass tissues, treated with a foliar application of potassium phosphite were subject to
High Performance Ion Chromatography (HPIC) analysis to determine PO33- and PO43(phosphate) amounts in leaf, root and crowns. It was determined that phosphite is rapidly
absorbed through the leaf tissues and readily translocated through the plants vascular system,
there were no significant increases in the PO43- levels in planta indicating that PO33- is not
readily metabolised to usable forms of P.
John Dempsey
HPIC analysis of phosphite treated turfgrass
Methodology
Treatments
Agrostis stolonifera, established in PVC growth vessels, filled with USGA specification
rootzone sand, were subject to foliar applications of potassium phosphite (KH2PO3). The
applications replicated standard turfgrass management rates of 0.35 g/m-2 of H3PO3.
Harvesting of the leaf and root tissues was carried out at pre-determined time periods: 1, 6,
12, 24, 48 hours and 1, 2, 3, 4, 5, and 6 weeks post application (p.a), crowns were harvested
at 4 and 6 weeks p.a., un-treated control plants provided a standard comparison.
Leaf tissues were collected using a scissors, at the set time periods, washed and rinsed in
distilled water, dried with tissue and then dried at 600C for 48 hours. Roots were collected by
placing the rootzone into a sieve and shaking to remove the soil. The roots were then washed
and dried as above. Crowns were harvested by firstly removing the leaf tissues and then
slicing the crowns away from the roots using a knife; they were then washed and dried as
above.
The dried tissues were finely ground and 0.5g were extracted into 10ml of water overnight,
passed through a 0.47 micron filter and injected into the sample loop of a Dionex HPIC
system using a 9mM sodium carbonate eluent. Results are reported as parts per million (ppm)
of dry tissue weight.
John Dempsey
HPIC analysis of phosphite treated turfgrass
Results
PO33- accumulation
Table 1 shows the data from the HPIC analysis and from these data figures 1, 2 and 3 were
configured. The data show that the highest rate of PO33- accumulations in the leaf tissues
were 48 h p.a. with a figure of 4889ppm, accumulations of 639ppm (13% of maximum
accumulation) 1 h p.a. and 3193ppm (65%) 6 h p.a were also determined. Two weeks p.a
accumulations in the leaf were at 2561 ppm, approximately 50% of the maximum and
gradually declined to 393 ppm at 6 weeks p.a.
Table 1 Data from HPIC analysis –sample weight, mg extracted and ppm for each sample.
Sample No and
Label
ppm
PO3 in
sample
ppm
PO4 in
sample
Sample
weight
gm.
ppm PO3
extract
ppm
P04
extract
1-leaf - control
2-Root - control
3-Leaf + 1 hour
4-Root + 1 hour
5-Leaf + 6 hours
6-Root + 6 hours
7-Leaf + 12 hours
8-Root + 12 hours
9-Leaf + 24 hours
10-Root + 24 hours
11-Leaf + 48 hours
12-Root + 48 hours
13-Leaf + 1 week
14-Root + 1 week
15-Leaf + 2 weeks
16-Root + 2 weeks
17-Leaf + 4 weeks
18-Root + 4 weeks
19-Crown + 4 weeks
20-Leaf + 6 weeks
21-Root + 6 weeks
22-Crown + 6 weeks
0
0
639
55
3193
111
3876
120
3608
376
4889
116
3334
126
2561
492
715
338
1250
393
65
484
13154
1435
6938
1197
7293
408
8499
1464
7165
1612
8590
1108
8229
937
10436
1389
5067
1658
4208
11124
449
4584
0.552
0.543
0.544
0.530
0.512
0.534
0.519
0.508
0.548
0.505
0.506
0.531
0.545
0.468
0.512
0.503
0.518
0.504
0.520
0.501
0.510
0.505
0.000
0.000
34.790
2.911
163.600
5.906
201.100
6.086
197.600
19.010
247.600
6.140
181.700
5.880
131.230
24.730
37.050
17.020
65.000
19.710
3.320
24.460
726.500
77.860
377.700
63.510
373.700
21.770
441.000
74.290
392.400
81.420
435.000
58.820
448.500
43.840
534.740
69.780
262.490
83.500
218.860
557.740
22.890
231.690
John Dempsey
mg
mg
extracted extracted
PO3
PO4
(10ml)
(10ml)
0.000
7.265
0.000
0.779
0.348
3.777
0.029
0.635
1.636
3.737
0.059
0.218
2.011
4.410
0.061
0.743
1.976
3.924
0.190
0.814
2.476
4.350
0.061
0.588
1.817
4.485
0.059
0.438
1.312
5.347
0.247
0.698
0.371
2.625
0.170
0.835
0.650
2.189
0.197
5.577
0.033
0.229
0.245
2.317
HPIC analysis of phosphite treated turfgrass
In the roots, accumulations were less than in the leaf tissues, with the highest rate of 492
ppm, achieved 2 weeks p.a, and accumulations of 55ppm (11%) 1 h p.a. and 111ppm (23%) 6
h p.a.
Analysis of the crowns from 4 and six weeks p.a showed levels of 1250 and 484 ppm
respectively.
Figure 1 PO33- accumulations in leaf and root tissues of A. stolonifera
John Dempsey
HPIC analysis of phosphite treated turfgrass
PO43- accumulation
PO43- levels had a mean value of 8092 ppm for leaf and 1136 ppm roots, which are within the
standard recommended levels for A. stolonifera. During the six week period of these analyses
the PO43- levels did not vary significantly from the mean value, indicating no in planta
conversion of PO33- to PO43-, figure 3 shows the combined PO33- and PO43- accumulations
over the six week period.
Figure 2 PO43- accumulations in leaf and root tissues of A. stolonifera.
John Dempsey
HPIC analysis of phosphite treated turfgrass
Figure 3 PO33- and PO43- accumulations in A. stolonifera over six weeks post application.
John Dempsey
HPIC analysis of phosphite treated turfgrass
Conclusions and further reseach
The significant points from these analyses are, firstly- phosphite is rapidly assimilated into
the leaf tissue of turfgrass and is translocated both in the xylem and phloem demonstrating
symplastic ambimobility within the tissues. The second point of interest is that there was no
significant increase in the PO43- levels, indicative that there is no rapid conversion of
phosphite to phosphate within the plant.
Even though the treatment and analyses were carried out during February, a period of low
growth and metabolism, assimilation into the leaf tissues was rapid, achieving 65% of the
maximum amount within 6 hours of application –a significant point for turfgrass managers; it
could be assumed that assimilation could be more rapid during higher growth conditions. The
level of phosphite within the leaf began to reduce after 48 hours and at 2 weeks p.a. had
dropped to 50% of the maximum accumulation, as most management programs apply
phosphite on a 2 to 3 week cycle this would indicate that levels would remain within the
range of between 2500 to 5000 ppm throughout the term of the program.
Further research is required in this area, the above study was carried out on greenhouse
samples with four replications for each analysis, which may have allowed some anomalies in
the data, and field studies have now begun using a 1,200 m2 turf nursery, which will allow for
a wide range of sample collection, increasing the validity of the data. These field studies will
gather data to hopefully answer a number of unresolved issuesData relating to the in planta fate of PO33- for example, over a longer application period, six
months to a year, does continuous treatment with phosphite lead to a cumulative increase in
the tissue levels and where in the plant does phosphite accumulate – within the cells or
extracellular spaces, is there a build up over time in meristematic areas, is there any in planta
metabolisation to other forms of P over a longer period than the six weeks of first analyses.
John Dempsey
HPIC analysis of phosphite treated turfgrass
References
Beard, J. and Oshikazu, T. (1997). Colour Atlas of Turfgrass Diseases. . New Jersey, Wiley
and Sons.
Cook, J., Landschoot, P. J. and Schlossberg, M. J. (2006). "Phosphonate products for
disease control and putting green quality." Golf Course Management: 93-96.
Cook, P. J., Landschoot, P. J. and Schlossberg, M. J. (2009). "Inhibition of Pythium spp.
and Suppression of Pythium Blight of Turfgrasses with Phosphonate Fungicides." Plant
Disease 93(8): 809-814.
Guest, D. and Grant, B. (1991). "The Complex Action of Phosphonates as Antifungal
Agents." Biological Reviews 66(2): 159-187.
Horvath, B. J., Mccall, D. S., Ervin, E. H. and Zhang, X. (2006). "Physiological Effects of
Phosphite Formulations on Turfgrass Challenged with Pythium and Heat Stress. Year II.".
Landschoot, P. J. and Cook, J. (2005). "Sorting out the phosphonate products." Golf Course
Management: 73-77.
Ouimette, D. G. and Coffey, M. D. (1988). "Quantitatave analysis of organic phosphonates,
Phosphonate, and other Inorganic Anions in Plants and Soil by Using High-Performance Ion
Chromatography." Phytopathology 78(9): 1150-1155.
Rickard, D. A. (2000). "Review of phosphorus acid and its salts as fertilizer materials."
Journal of Plant Nutrition 23(2): 161 - 180.
Sanders, P. L. (1983). "Control of Pythium spp. and Pythium Blight of Turfgrass with
Fosetyl Aluminum." Plant Disease 67(12): 1382-1383.
Schroetter, S., Angeles-Wedler, D., Kreuzig, R. and Schnug, E. (2006). "Effects of
phosphite on phosphorus supply in corn (Zea mays)." Landbauforschung Volkenrode 56: 8799.
Vincelli, P. and Dixon, E. (2005). "Performance of selected phosphite fungicides on greens."
Golf Course Management.
John Dempsey