Taxus and Taxol - A Compilation of Research Findings

Special Circular 150-99

Growth Rate of Container-Grown Taxus x media 'Hicksii' and 'Densiformis' Compared at Two Levels of Nutrition and Irrigation

Robert C. Hansen,
Department of Food, Agricultural, and Biological Engineering,
Ohio Agricultural Research and Development Center,
The Ohio State University,
Wooster, Ohio.

Kenneth D. Cochran,
Secrest Arboretum,
Ohio Agricultural Research and Development Center,
The Ohio State University,
and The Ohio State University Agricultural Technical Institute,
Wooster, Ohio.

R. Peter Fynn,
Department of Food, Agricultural, and Biological Engineering,
Ohio Agricultural Research and Development Center,
The Ohio State University,
Wooster, Ohio.

Introduction

The effect of fertilization on the growth of Taxus cultivars and other woody ornamental species has been studied previously. Flint (1962) defined the upper limit of soil nitrogen for Taxus x media 'densiformis' along with four other woody ornamentals, i.e., Forsythia intermedia 'Lynwood Gold,' Viburnum plicatum var. tomentosum, Rhododendron catawbiense 'grandiflorum,' and Pieris japonica. The rooted cuttings and Pieris japonica seedlings were planted in 15 cm containers using standard potting medium. N was supplied by injecting soluble concentrates in the irrigation water. N levels of 100, 200, 300, and 400 ppm were compared while the concentration of K in the irrigation water was held constant at 200 ppm. This resulted in mean levels of 10, 19, 31, and 41 ppm N, respectively, and 19 ppm K in the soil extract. Growth was measured by oven drying detached plant tops 100 to 110 days after planting. The upper limit of N for optimum growth was between 10 and 19 ppm for Viburnum and Rhododendron and between 19 and 31 ppm for Pieris. Meanwhile, the upper limit of N for optimum growth was between 31 and 41 ppm in soil extract for Taxus x media 'densiformis' and Forsythia.

Gouin and Link (1966) studied the effects of three levels each (27 different combinations) of N, P, and K on total growth and nutrient levels in the foliage of Taxus x media 'Hatfieldii.' Rooted cuttings were planted in 25 cm pots in a standard medium and grown for 19 months after which plant tops were pruned to the top of the pots and dried. Maximum total growth occurred when levels of N, P, and K were 224, 75, and 135 ppm, respectively. According to Gouin and Link (1996): "Any variation from these levels not only resulted in significant decrease in total growth, but also caused drastic changes in the nutrient content of the foliage." They concluded that Taxus growth was strongly affected by any deviation from a nutritional balance of 3-1-2 for N-P-K, respectively.

Meyer and Tukey (1967) showed that the timing of nutrient applications and root temperature have important effects on root growth of Taxus x media 'Hicksii' and Forsythia intermedia 'Spring Glory.' Roots of both species grew appreciably over winter, while the above-ground parts were dormant. The average root growth of plants receiving nutrient applications was much greater than that of plants receiving none. In addition, increase in nutrient reserves during the dormant season increased the amount of shoot growth during the following spring.

While research has been targeted to identify nutritional and cultural practices that maximize the production of Taxus biomass, no recommendations have been reported for sustaining and maximizing annual production of clippings while the plant continues to grow year after year. Also, which nutritional and cultural practices lead to optimum concentration of Taxol in clippings? The research reported here was a first attempt to explore these issues.

Objective

The objective of the experiment was to measure the effect of Taxus cultivar, nutrition level, and irrigation frequency on the total mass of Taxus clippings produced and on the Taxol concentration in the clippings.

Materials and Methods

Experiments were conducted June 1, 1992, to September 15, 1993, in a double-polyethylene greenhouse located at the Ohio Agricultural Research and Development Center (OARDC), Wooster, Ohio (41°30' N Latitude). Greenhouse air temperatures ranged from 20 to 40°C. The specific greenhouse compartment used in this experiment was 5 m wide x 30 m long with an evaporative pad and fan ventilation system across the short dimension.

The plants were potted June 15, 1992, in 22 cm (8.5 in./#2) containers using 60% pine bark, 30% Sphagnum peat moss, and 10% Haydite®. The mix also included 3 kg/m³ (5 lbs/yd³) of lime. The plants were placed on a dirt floor covered with a weed mat made of black fabric. Supplemental lighting was supplied with HID lights December 14, 1992, through April 5, 1993, in an attempt to avoid plant dormancy. The lights were set to come on at 7 a.m. and were shut off at 10 p.m.

The experiment was designed to compare two cultivars - Taxus x media 'Hicksii' and Taxus x media 'Densiformis.' Two hundred plants were obtained from Zelenka Nursery, Inc., Grand Haven, Michigan - 100 'Hicksii' as two-year liners and 100 'Densiformis' as three-year liners. Half of the plants in each cultivar were irrigated two times per week [Monday and Friday (M-F)] while the other half were irrigated four times per week [Monday, Wednesday, Friday, and Sunday (M-W-F-Sun)].

Finally, the plants were supplied with nitrogen, phosphorus, and potassium from soluble concentrates (21-7-7) in the irrigation water. Approximately 100 ppm N, 33 ppm P, and 33 ppm K were delivered to half of the plants as the low level of nutrition compared to 200 ppm N, 66 ppm P, and 66 ppm K as the high level for the other half. Appropriate levels of soluble micronutrient concentrates were included in each solution.

Tables 1 and 2 summarize the factors and levels used in the full factorial, three-factor, two-level experimental design. The design resulted in eight treatments where each treatment was replicated four times using five plants per replication. Therefore, 80 'Hicksii' and 80 'Densiformis' were included in the experiment. An additional 52 plants were used as border plants.

Figure 1 is a schematic drawing showing the randomized location of each treatment within four replications. The drawing also illustrates the layout for four irrigation lines that were required to deliver high and low levels of nutrition using the two irrigation schedules referred to earlier. Water and nutrients were delivered through 16 mm polyethylene tubing used as a main line. Water was delivered to each individual plant through 3 mm spaghetti tubing using 0.03 L/min (0.5 gph) woodpecker drippers. Each irrigation event required 39 minutes on average before water dripped through the bottom of the pots. During this time period, approximately 760 ml of water along with designated nutrient levels were delivered to each pot. Automatic timers were used to set the days of the week and the duration for each irrigation.

A Q-COM (Q-COM Corporation, Irvine, CA 92614) moisture tension measurement system was used to monitor tension in one container per treatment. Each tensiometer was placed half way down into the growing medium of each of the four containers. These tensiometers used a "high flow" ceramic cup for the tip and had a relatively fast reaction time to changes in moisture tension. The tension readings typically ranged between 2 kPa and 10 kPa throughout the study. This particular regimen seemed to maintain good soil moisture conditions while avoiding water drainage from the bottom of the containers.

Growth rate was measured based on plant volume increase and weight of clippings collected after a typical pruning event. Plant volume was represented with a pseudo cylinder by averaging two diametrical measurements of each plant, calculating a projected area, and then multiplying by plant height. Measured diameters were approximately at right angles to each other. Height was defined by the distance from a plane defined by the top of the pot to the furthermost extent of branching in the vertical direction. Each diameter and height dimension was defined with a caliper. The quantity of clippings harvested and weighed was determined by subjectively simulating what a grower would typically remove during an annual pruning event to bring about compactness and an aesthetically pleasing shape.

Measurements of plant volume were done on one randomly selected plant per treatment per replication, i.e., eight treatments times four replications equals 32 plants. Plant volumes were first measured 8/4/92. Thereafter, measurements were made 10/4/92, 11/4/92, 1/12/93, 2/23/93, and 5/4/93 and subtracted from 8/4/92 results to determine volume increase. All 80 'Densiformis' plants were pruned 7/8/93, while all 80 'Hicksii' plants were pruned 9/8/93. The clippings were collected, immediately placed in one-gallon plastic zip-lock bags, and weighed as harvested without drying.

Figure 1. Top view of the experimental layout showing random placement
of eight treatments and associated irrigation lines within four replications.

Results and Discussion

Since the plants were potted in June, the greatest flush of growth for Taxus cultivars was expected during late summer, between July and October. A summary of growth in terms of volume increase for each treatment is shown in Table 3 for the two-month period (8/4/92 to 10/4/92). Each measurement is the average of four replications of randomly selected plants. The average growth for the 'Densiformis' samples was nearly five times as great as the growth measured for 'Hicksii' (2,109 vs. 425 m³/month). Treatment No. 7 consisting of 'Densiformis' irrigated four times per week (M-W-F-Sun) using 100-33-33 ppm N-P-K resulted in the greatest growth of all treatments with 2,534 cm³/month on average while 'Hicksii' irrigated two days per week (M-F) using 100-33-33 ppm N-P-K resulted in the least volume increase with 219 cm³/month. The effect of growing 'Densiformis' using the low nutrition level of 100-33-33 ppm N-P-K (Treatment No. 5 and 7) averaged 2,471 cm³/month while the high level of nutrition 200-66-66 ppm N-P-K (Treatment No. 6 and 8) averaged only 1,747 cm³/month (Table 3). Finally, there was little difference in volume increase for 'Densiformis' when irrigating two times per week (2,029 cm³/month) vs. four times per week (2,189 cm³/month). Based on these very preliminary results using data from the two-month period (8/4/92 to 10/4/92), the best combination of factors and levels for maximum growth was 'Densiformis' irrigated two times per week using the low nutrition level 100-33-33 ppm N-P-K.

Table 4 shows example diameter and height measurements along with calculated volume for one of the largest 'Densiformis' plants and one of the largest 'Hicksii' plants, 8/4/92 through 5/4/93. Even though HID lighting was used over winter, the results show that little growth occurred and that a period of dormancy was not avoided. Decreases in volume (e.g., 'Densiformis,' Nov. 4 to Jan. 12; 'Hicksii,' Oct. 4 to Jan. 12) would imply plants diminished in size somehow. Since no pruning was done, the results most probably indicate variation in the technician's ability to subjectively identify representative diameters month to month. Note that the 'Densiformis' plant increased in volume from 6,509 to 16,604 cm³/month (larger by a factor of 2.55, 8/4/92 through 5/4/93) while the 'Hicksii' increased from 1,862 to 5,025 cm³/month (larger by a factor of 2.70, 8/4/92 through 5/4/93). Because of the preliminary nature of this research, the impact of dormancy and undesirable environmental conditions in the greenhouse throughout the winter, results obtained for volume increase after 10/4/92 through 5/4/93 for the rest of the plants did not merit further analysis.

A preliminary analysis of expected clippings mass that might be harvested from Taxus x media 'Hicksii' and Taxus x media 'Densiformis' is shown in Table 5. No clippings were harvested until near the end of the experiment. 'Densiformis' was pruned 7/8/93 and 'Hicksii' was pruned two months later, 9/8/93, i.e., the 'Hicksii' grew for 15 months after potting and the 'Densiformis' grew for only 13 months before the clippings were harvested. Based on the results for increase in volume (Table 3), the clippings mass for 'Densiformis' was expected to be much greater than for 'Hicksii.' Instead, the average yield for the 80 'Hicksii' plants was 28 grams per plant compared to 26 grams per plant on average for the 80 'Densiformis' plants. The plant volume represented by the pseudo cylinder clearly did not correlate with mass of clippings harvested. 'Densiformis' clippings did appear to be more delicate (smaller stem diameter than 'Hicksii') and more open and widely dispersed in the pseudo cylinder than 'Hicksii.' 'Hicksii' was typically more compact. This observation might partially explain the unexpected results. Based on an appropriate statistical analysis of variance, no significant difference was discerned between the eight treatments for the clippings mass response variable. This was due in part to the large standard deviation found within the treatment results.

As a comparison, Hansen et al. (1994) reported clippings yields for three-year 'Densiformis' were 23.6 grams per plant (dry basis) while yields for three-year 'Hicksii' were 18.8 grams per plant (dry basis). These measurements were obtained from field-grown plants at commercial nursery operations. Typical moisture contents for clippings were 51 to 57% (wb).

Conclusions

• Growth in terms of volume increase was measured for the two-month period (8/4/92 to 10/4/92). The average growth for the 'Densiformis' samples was nearly five times as great as the growth measured for 'Hicksii' (2,109 vs. 425 cm³/month).
• Treatment No. 7, consisting of 'Densiformis' irrigated four times per week (M-W-F-Sun) using 100-33-33 ppm N-P-K, resulted in the greatest growth of all treatments with 2,534 cm³/month on average while 'Hicksii' irrigated two days per week (M-F) using 100-33-33 ppm N-P-K resulted in the least volume increase with 219 cm³/month.
• The effect of growing 'Densiformis' using the low nutrition level of 100-33-33 ppm N-P-K (Treatment No. 5 and 7) averaged 2,471 cm³/month while the high level of nutrition 200-66-66 ppm N-P-K (Treatment No. 6 and 8) averaged only 1,747 cm³/month.
• One of the best 'Densiformis' plants increased in volume from 6,509 to 16,604 cm³/month (larger by a factor of 2.55, 8/4/92 through 5/4/93) while the best 'Hicksii' increased from 1,862 to 5,025 cm³/month (larger by a factor of 2.70, 8/4/92 through 5/4/93).
• Based on harvested clippings mass (wb), the average yield for 80 'Hicksii' plants was 28 grams per plant compared to 26 grams per plant on average for 80 'Densiformis' plants.

Sources of Error

Taxus cultivars are typically grown outdoors in soil rather than in a greenhouse in potting medium. The effect of growing these plants in an artificial environment without full spectrum lighting is not known. Also, the attempt to provide HID lighting during the winter months to avoid dormancy is questionable. Since dormancy occurred anyway and growth was minimal, the plants in treatments set for irrigation four times per week (M-W-F-Sun) were no doubt watered in excess. In addition, the continual addition of nutrients during dormancy may have led to a build up of salts in the potting media, leading to detrimental results. A final difficulty was encountered when attempting to monitor growth by measuring plant diameter and height. It was difficult to determine actual dimensions to be measured. Because the choice was very subjective, measurements varied significantly between technicians and from month to month with the same technician.

Some of the plants developed rust-colored leaves/needles by May of 1993. The needles would fall off when the plant was mildly shaken. A plant pathologist thought the symptom might be due to salt build up in the potting medium. He did not recognize any disease symptoms. Since new growth was evident, the pathologist was convinced the plants were not dying. Original objectives for the experiment included measurement of the Taxol content of harvested clippings. Because of the experimental difficulties and associated sources of error noted in the two earlier paragraphs and because of the cost of conducting laboratory analyses for Taxol, these measurements did not seem to be justified and therefore were not done.

References

Flint, H. L. 1962. Nitrogen range for optimum growth of some woody ornamental species grown in containers. American Society for Horticultural Science 80: 622-624.

Gouin, F. R. and C. B. Link. 1966. The effects of various levels of nitrogen, phosphorus, and potassium on the growth and chemical composition of Taxus media cv 'Hatfieldi.' American Society for Horticultural Science 89: 702-705.

Hansen, R. C., K. D. Cochran, H. M. Keener, and E. M. Croom Jr. 1994. Taxus populations and clippings yields at commercial nurseries. HortTechnology 4(4): 372-376.

Meyer, M. M. and H. B. Tukey Jr. 1967. Influence of root temperature and nutrient applications on root growth and mineral nutrient content of Taxus and Forsythia plants during the dormant season. American Society for Horticultural Science 90: 440-446.