ABSTRACT
The effects of 50% or 100% of a herbage diet of plantain on milk production and urinary nitrogen (N) concentration were measured in two experiments for late (autumn 2015) and early (spring 2015) lactation dairy cows. Three groups of 12 mixed age Friesian × Jersey dairy cows were offered a perennial ryegrass-white clover pasture, or pure plantain or 50% perennial ryegrass-white clover and 50% pure plantain by ground area (50–50 pasture–plantain). Urine N concentration was lower in both experiments (P < .001) for plantain (2.4 and 2.2 g N/L) and 50–50 pasture–plantain (3.6 and 3.4 g N/L) than pasture (5.4 and 4.7 g N/L). Cows on plantain or 50–50 pasture-plantain produced at least as much milk as those on pasture in both experiments. Plantain may offer environmental benefits to dairy systems by reducing the N concentration of urine deposited on the soil from grazing cows.
Introduction
Reducing the environmental impact of the dairy industry in New Zealand has become a key focus with an increasing interest in water quality issues, in particular nitrogen (N) leaching from pastoral land (Bryant et al. Citation2007; Woodward et al. Citation2012). Regional Councils throughout New Zealand have been developing regulations that place a limit on the amount of nitrate-N leached from agricultural land. Nitrogen from urine patches is a major contributor to N leaching (Di and Cameron Citation2007). There is a large discrepancy between the N content of grazed forages, and the N requirement of the animals; N in excess of an animal’s requirement is excreted, primarily in the urine (Tamminga Citation1992). This leads to high N loading (up to 1000 kg N/ha) on the soil at the urine patch, which is easily leached when drainage occurs (Di and Cameron Citation2007). Mitigation strategies have utilised variations in the chemical composition (water-soluble carbohydrate (WSC) and crude protein (CP)), mineral profile and secondary plant compounds in forages, to reduce urinary N excretion or divert dietary N away from urine. Previous experiments (Woodward et al. Citation2012; Totty et al. Citation2013; Edwards et al. Citation2015; Bryant et al. Citation2017) achieved lower urinary N from cows by incorporating alternative pastures species including plantain (Plantago lanceolata), chicory (Cichorium intybus) and additional legumes such as red clover (Trifolium pratense) and lucerne (Medicago sativa) into pasture mixtures with perennial ryegrass (Lolium perenne) and white clover (Trifolium repens). In comparison with standard perennial ryegrass-white clover pastures, these diverse mixtures have shown consistent reductions in urinary N concentration of at least 20% while maintaining or improving milk production (Woodward et al. Citation2012; Totty et al. Citation2013; Bryant et al. Citation2017). However, it is unclear whether the effects are due to an individual species in the mixture rather than the diversity. Research using sheep fed plantain or perennial ryegrass diets indicated a positive relationship between plantain feeding and urine volume (O'Connell et al. Citation2016) which may explain the differences in urine N concentration observed in dairy cows. There is currently little research comparing perennial ryegrass pasture with plantain forage fed as a sole diet. The objective of this study was to assess whether plantain as a grazed pasture species can support milk production similar to that achieved with standard ryegrass-white clover while reducing urinary N losses. We hypothesise that cows grazing plantain or a 50–50 pasture–plantain diet will produce as much milk as those on pasture with lower urinary N concentrations.
Materials and methods
Experimental site and treatments
Two experiments were conducted at the Lincoln University Research Dairy Farm (LURDF) with the approval of the Lincoln University Animal Ethics Committee in late (AEC 592) and early (AEC 639) lactation in 2015. There were three pasture treatments: (1) a perennial ryegrass-white clover pasture (n = 12 cows); (2) plantain (n = 12 cows) and (3) an area that was made of 50% perennial ryegrass-white clover and 50% pure plantain by ground area (n = 12 cows). A group of 36 pregnant, lactating Friesian × Jersey dairy cows were blocked into three groups of 12 cows according to milk solids production (1.31 ± 0.03 kg MS/cow/day for late and 2.48 ± 0.04 kg MS/cow/day for early lactation), age (5.9 ± 0.4 years for late and 5.6 ± 0.3 years for the early lactation experiment), days in milk (217 ± 3.2 days for the late and 50.9 ± 1.1 days for the early lactation experiments) and liveweight (515 ± 8.6 kg for the late and 496 ± 8.4 kg for the early lactation experiments) (all mean ± SEM). Both studies were conducted over a 10-day period with late lactation occurring from 19 to 29 March 2015 and the early lactation experiment occurring from 8 to 18 October 2015. This included a 3-day transition period where cows grazed their respective pasture treatments with no measurements and a 7-day experimental period. Before the experiments, all cows grazed perennial ryegrass-white clover pasture together. Cows were milked twice daily at approximately 0600 and 1400 h.
The perennial ryegrass (cv. Arrow)-white clover (cv. Kopu II) pastures were established across one-half of three 1.5 ha paddocks in March 2014. In December 2014, the remaining half of the same three paddocks were sown in plantain (cv. Tonic). Paddocks were grazed with dairy cows rotationally to a constant height of approximately 1500 kg DM/ha and fertilised with urea at a rate of 70 kg N/ha 25 days prior to the experiment. Pasture was irrigated with a centre-pivot irrigator. Each 1.5 ha paddock was separated into the three forage treatments, of pasture only, a 50–50 pasture–plantain and pure plantain area, using electric fencing. Due to low pasture growth rates in the late lactation study, more area was required to meet feed allocation targets so an additional perennial ryegrass and white clover pasture paddock was included for the 50–50 pasture–plantain treatment.
After each morning milking, cows were offered a target daily allowance of 18 kg DM/cow/day above 3.5 cm in the late lactation experiment and 30 kg DM/cow/day above ground level in the early lactation experiment. Daily herbage allocation during the experiment was based on a national calibration equation for perennial ryegrass-white clover (kg DM/ha = 140 × Rising Plate Meter (RPM) reading + 150) and previously derived calibration equations between herbage mass and pasture height for plantain pastures (kg DM/ha = 94 × RPM reading + 455) (Haultain et al. Citation2014). For the 50–50 pasture–plantain treatment pre-grazing plate meter readings were taken in both the pasture and plantain areas. The sum of 0.5 × pasture herbage mass and 0.5 × plantain herbage mass was used to determine the area to be grazed as spatially separated pasture and plantain. Daily allocated areas were controlled by temporary electric fencing. Back fencing was used to prevent grazing of residual regrowth. Cows had ad lib access to water through a portable trough.
Herbage measurements
At least 50 compressed pasture height measurements were recorded daily pre- and post-grazing using a calibrated rising plate meter (RPM; Jenquip, Filip’s EC 09, Electronic Folding plate meter). The pre-grazing measurements were recorded in the area estimated to be allocated in the next forage allocation. Calibration measurements were collected from pastures every second day by cutting two 0.2 m2 quadrats to ground level in late lactation and five in early lactation before and after grazing during each experiment. Two RPM measurements were recorded in each quadrat prior to harvesting herbage. Collected herbage samples were weighed fresh and a subsample taken to determine botanical composition of pastures. All botanical components and bulk samples were oven dried at 65°C for 48 h and total dry weight and DM% determined. Linear and curvilinear relationships between herbage mass and pasture height were compared and best-fit equations (greatest r2) were fitted to the data. The calibration equations for each pasture type were: ryegrass-clover (kg DM/ha) = 145 × height (RPM clicks) + 3.92, r2 = 0.85 and plantain (kg DM/ha) = 73 × height (RPM clicks) + 248, r2 = 0.89. In the late lactation experiment, calibration equations were: pasture (kg DM/ha) = 120 × height (RPM clicks) + 312, r2 = 0.63 and plantain (kg DM/ha) = 114 × height (RPM clicks) + 470, r2 = 0.71. Using equations derived from experimental data sets and grazing areas, the actual daily herbage allocation was calculated as 21.4 ± 2.4 kg/DM/cow for pasture, 19.1 ± 0.94 kg/DM/cow for plantain and 22.2 ± 0.9 kg/DM/cow for 50–50 pasture–plantain in late lactation. In the early lactation experiment, actual daily herbage allocation was calculated as 32.8 ± 1.3 kg/DM/cow for pasture, 31.3 ± 0.65 kg/DM/cow for plantain and 31.6 ± 0.37 kg/DM/cow for 50–50 pasture–plantain. Apparent group DM intake of cows was calculated from herbage disappearance between pre- and post-grazing calibrated RPM measurements and areas allocated.
Subsampled herbage was manually sorted into sown species, weeds and dead material. Sorted material and a bulk sample of the pasture was oven dried at 65°C for 48 h to determine botanical composition of pastures. Bulk oven-dried samples were ground through a 1-mm sieve and scanned by a near infra-red spectrophotometer (NIRS, NIRSystems 5000, Foss, Maryland, USA) to determine CP, digestible organic matter in the dry matter (DOMD), WSC (including soluble starch), acid detergent fibre (ADF) and neutral-detergent fibre (NDF). Calibration equations for NIRS were derived from perennial ryegrass, clover and plantain herbage. Ground samples were then subsampled and analysed for mineral composition by inductively coupled plasma atomic emission spectroscopy. Plantain samples were also analysed for secondary plant compounds (catalpol, aucubin and acteoside) using high performance liquid chromatography by Massey University analytical laboratory following procedures outlined by Tamura and Nibishe (Citation2002). Metabolisable energy (ME) was calculated as MJ ME/kg DM = 0.16 × DOMD (CSIRO Citation2007).
Animal measurements
Milk yield was measured daily for individual cows with an automated system (DeLavel Alpro Herd Management System, DeLavel, Tumba, Sweden). Milk samples from all cows were collected on days 4, 6, 8 and 10 from the afternoon and following morning milking to determine milk composition and milk urea nitrogen (MUN). Fat, protein and lactose composition in milk were analysed by Livestock Improvement Corporation Ltd (Christchurch, New Zealand) by MilkoScan (Foss Electric, Hillerod, Denmark). Samples for MUN were centrifuged at 4000g for 10 min at room temperature and refrigerated for 10 min to allow the fat to solidify on the top and be removed. Skim milk was pipetted into a clean micro-centrifuge tube, and frozen for later analysis of thawed samples using an automated Modular P analyser (Roche/Hitachi).
Spot samples of urine and faeces were collected on days 5, 7 and 9 of the experiments from all cows following morning and afternoon milking. Spot urine samples were collected mid-stream by vulval stimulation and were immediately acidified with hydrochloric acid to prevent volatilisation and stored at −20°C. Faeces were obtained from voluntary events in the yards or by rectal stimulation and stored at −20°C. Faeces were freeze dried and ground through a 1-mm sieve. Faecal N content was determined by combustion under oxygen (Elemental Analyser vario MAX CN, Analysensysteme GmbH, Hanau, Germany). Urine N content of thawed subsamples was determined by an autoanalyser (Roche Cobas Mira Plus CC, Minnesota, USA). Urea content of acidified urine was analysed using a commercial enzymatic kinetic technique (Randox, Crumlin, Co., Antrim, UK). Creatinine content of acidified urine was determined calorimetrically using a commercial kit from Randox laboratories (Crumlin, Co., Antrim, UK).
A urine meter harness was worn by all cows for up to 24 h between days 5 and 10 for both experiments. The harness involved attaching a flow meter to the vulva of the cow which transmits pulse signals, whenever the cow urinates, to a data logger carried in a cover worn by the cow (Ravera et al. Citation2015). On occasions, the urine meter would fail through becoming unstuck, wire breakage or sensor failure. This left 20 cows in the late lactation experiment (six from pasture, seven from 50–50 pasture–plantain and seven from plantain) from which data were used and 22 cows in the early lactation experiment (eight from pasture, seven from 50–50 pasture–plantain and seven from plantain).
Urinary N output was estimated using three methods: (1) urine volumes measured using the urine harness were multiplied by overall treatment urine N concentration averages, (2) creatinine-based equation: urinary N (g/day) = (21.9 (mg/kg) × LW (kg) × (1/urinary creatinine (mg/kg))) × urine N (g/kg) (Pacheco et al. Citation2007) and (3) MUN-based equation: urinary N (g/day) = 0.026 × LW (kg) × milk urea N (mg/dL) (Jonker et al. Citation1998).
The effect of treatment on rumen microbial activity was calculated from the ratio of purine derivatives (PD) to creatinine in spot samples of urine (Chen Citation1989; Verbic et al. Citation1990; Chen et al. Citation1995). The resulting PD index is a relative measure of microbial protein synthesis:where total PD [allantoin (mmol/L) + uric acid (mmol/L)].
Statistical analysis
Treatment means for milk, urine and faeces were determined using data from individual measurements from animal samples over sampling days. The effect of pasture type on milk, urine and faecal measurements was analysed for variance using GenStat 15.1, with cows as random effect and pasture type as fixed effect using a one-way ANOVA. Results were declared to be significant at P < .05. Means were separated using Fisher’s protected least significance difference test. No statistical analysis was carried out on herbage samples or apparent DM intake as they were based on samples collected from each daily allocation within the same paddock. Intake and herbage measurements were estimated as means for the treatment group as animals grazed together in their treatment groups.
Results
Herbage
Herbage characteristics and composition of plantain and ryegrass-white clover pastures are shown in . In the late lactation experiment, pre-grazing mass of the pasture portion of the 50–50 pasture–plantain was 700 kg DM/ha greater than the herbage mass for any other treatment. Dry matter content of plantain was nearly half that of ryegrass in the late lactation experiment and about 20% lower in the early lactation experiment. Organic matter content was lower in plantain than in ryegrass in the late lactation experiment only. The mineral composition of pasture and plantain is shown in .
Table 1. Mean herbage characteristics and chemical composition (±SEM) of pasture, plantain or 50–50 pasture–plantain sampled to ground level.
Table 2. Apparent intake of dairy cows grazing pasture, plantain or 50–50 pasture–plantain in autumn and spring and water intake in spring (±SEM).
The post-grazing mass of plantain appeared to be lower than that of pasture in both experiments (). The apparent DM intake increased by about 20% for cows grazing plantain and 50–50 pasture–plantain compared with those grazing pasture (). In the early lactation experiment, apparent DM intake was greater for the plantain treatment only. The ME and CP was similar across pasture treatments for both experiments (). Similarly, apparent N intake and ME intake did not differ across treatments.
Table 3. Mean herbage mineral composition (% DM) bioactive glycoside (mg/g dry DM) (±SEM) of pasture, plantain or 50–50 pasture–plantain sampled to ground level.
Total average water intake was similar for all treatments (150 L/cow/day) and more than two-thirds of total water were ingested in feed. As plantain intake increased, water intake from feed increased and drinking water declined ().
The mineral concentration was similar for pasture and plantain except for sodium and calcium. Sodium concentration was two times greater in plantain than pasture in the late lactation experiment only. Calcium concentration of the herbage was about three times greater in plantain than pasture for both experiments. Of the secondary plant compounds measured, acteoside had the highest concentration in plantain (). Catalpol was present in very low concentrations (<0.09%). The concentration of all secondary compounds in plantain tended to be higher in the late lactation experiment.
Milk production and composition
Milk volume (L) and milk solids yield was greatest (P < .05) for pure plantain than ryegrass pasture in the late lactation experiment though there were no treatment differences in the early lactation experiment (). In both experiments, ryegrass and plantain (50–50) altered milk composition. In the late lactation experiment, milk fat percent was lower and lactose percent was higher when cows grazed 50–50 pasture–plantain compared with ryegrass pasture alone (). In the early lactation experiment, milk protein percent was lower for the 50–50 treatment than the ryegrass pasture treatment. Milk urea N was lowest (P < .005) for plantain in both experiments.
Table 4. Mean milk yield and composition (n = 12) of dairy cows grazing perennial ryegrass-white clover pasture, plantain or 50–50 pasture–plantain.
Urine and faeces
The concentration of urea and N in urine was lowest (P < .001) for plantain, intermediate for 50–50 pasture–plantain and highest for pasture in both experiments (). In the late lactation experiment, the concentration of NH3 in the urine was lowest for plantain. There was no difference in NH3 in the urine among treatments in the early lactation experiment. Estimated urinary N output was lowest for plantain for all methods used in both experiments ().
Table 5. Mean urine and faecal N characteristics (n = 12) and urine volume and urination frequency of dairy cows grazing perennial ryegrass-white clover pasture, plantain or 50–50 pasture–plantain.
In the late lactation experiment, there was no difference in the average urination frequency or volume between treatment groups as measured by the urine harness (). Using measured total urine volume per cow per day, daily N output was calculated to be 30% lower for cows on plantain (177 g N/cow/day) than those grazing pasture (251 g N/cow/day) in the late lactation experiment. In the early lactation experiment, daily N output was calculated to be over 40% lower for cows on plantain (119 g N/cow/day) and 50–50 pasture–plantain (114 g N/cow/day) than those on pasture (205 g N/cow/day)
The concentration of N in the faeces was similar among treatments in the late lactation experiment.
Microbial protein supply
Urinary PD concentrations and calculations associated with estimates of microbial N supply are presented in . The concentration of PD’s allantoin, uric acid and hippuric acid was lowest (P < .001) for plantain in both experiments. All PD concentrations tended to be higher in the early lactation experiment than in the late lactation experiment for all treatments.
Table 6. Mean urine purine derivative (PD) concentrations (mmol/L) of dairy cows (n = 12) grazing perennial ryegrass-white clover pasture, plantain or 50–50 pasture–plantain.
Discussion
The hypothesis of the present study was that the use of plantain as a grazed pasture species would be suitable to achieve similar milk production to that achieved by feeding standard ryegrass-white clover, while reducing urinary N losses. The results from this study support this hypothesis. In these experiments, pure plantain swards as 100% or 50% of a cow’s forage allowance reduced the concentration of N in the urine and estimated N excretion per day while maintaining or improving milk production.
Milk production
Milk solids produced per cow in late lactation was greatest for cows grazing plantain (1.67 kg/day) compared to pasture (1.50 kg/day) with 50–50 pasture–plantain intermediate (1.60 kg/day). This was largely due to an increase in milk volume as milk protein percentage was unaffected by pasture treatment and fat percentage tended to be lower where plantain was included in the diet. An overall increase in milk solids and a reduction in milk fat were also observed by Totty et al. (Citation2013) when plantain was included in a mixed pasture. A possible explanation for the increase in milk volume was the observed increase in apparent dry matter intake (DMI) for cows on plantain in the late lactation experiment. This occurred because plantain was grazed to a lower post-grazing herbage mass compared with pasture despite similar daily allocations of herbage. However, it is noteworthy that cows grazing 50–50 pasture–plantain had the highest apparent DMI in late lactation and this did not translate to increased milk production. In the early lactation experiment, milk production was not different between pasture types, despite some differences in apparent DMI. In a related study, Edwards et al. (Citation2015) reported differences in DMI between simple and diverse pastures (15.3 ± 2.1 vs. 16.2 ± 1.4kg DM/cow/day, respectively) that did not change the milk yield and composition.
N partitioning
Previous short duration experiments have shown that when plantain is included in mixtures with ryegrass, white clover and chicory, urinary N concentration of dairy cows was reduced by at least 20% compared with perennial ryegrass-white clover pastures (Woodward et al. Citation2012; Totty et al. Citation2013). In the current experiment, when plantain was included in the diet at 50% or 100% of the allocation, urinary N concentration was more than 30% lower, in both experiments, compared with perennial ryegrass-white clover pasture only. The MUN concentration, an indicator of surplus dietary N (Cosgrove et al. Citation2014), was also lower for plantain-fed cows than for pasture-fed cows. Previous studies have shown the excretion of N in urine to be linearly related to N intake (Tas et al. Citation2006; Higgs et al. Citation2012). The current experiment indicated that factors other than N intake may have been a driver of lower urinary N concentration as there is no clear relationship with N intake across the dietary treatments. There is some evidence from in vitro studies that aucubin and acteoside may reduce rumen ammonia formation (Navarrete et al. Citation2016). There was no difference in faecal nitrogen, suggesting that this was not a route of increased excretion. Similar results have been seen by Judson and Edwards (Citation2016) where urinary N concentration was reduced by about 30% for heifers supplemented with plantain silage despite having similar N intakes to those with no plantain in their diet. There may have been some differences in energy and protein supply to the rumen indicated by the lower PD index for cows on plantain (Chen Citation1989).
The urinary N concentration can be influenced by total urine volume. The total volume of urine excreted per day by animals grazing plantain (74 L) in the late lactation experiment measured by the urine harness was 57% greater than the volume of urine excreted by cows grazing pasture (47 L). Further, creatinine, a marker of urine volume (Chizzotti et al. Citation2008; Waldrip et al. Citation2013), was lower for cows on plantain diets in both experiments, which suggested a higher urinary output in both experiments for diets containing plantain. This may have been a factor of higher mineral content, secondary plant compounds or increased water intake due to the lower DM% of plantain. Plantain in both the late and early lactation experiments had a greater calcium and sodium mineral content than pasture. The concentration of sodium, a known diuretic (Ledgard et al. Citation2015), was almost twice as much in plantain compared with pasture but only in the late lactation experiment. This coincided with an observed increase in urine volume indicating that sodium may have been a factor. This increase in urine volume was supported by recent data, which strongly suggest a sustained diuretic effect when plantain was fed to sheep (O’Connell et al. Citation2016). Although the authors did not measure any secondary compounds, O’Connell et al. (Citation2016) suggested that diuresis may have been due to bioactive compounds which are known to exist in the leaves of plantain.
Bioactive compounds in plantain have been studied in early medicinal work (Nishibi and Mural Citation1995). The major compounds in plantain include iridoid glucosides catalpol and aucubin and phenylethanoid glucoside acteoside. Although catalpol is a known diuretic (Tamura and Nishibe Citation2002), the availability of the compound in this experiment is unlikely to explain the higher urine volume output for cows grazing plantain as its concentration was low (<0.08 g/kg DM). The aucubin and acteoside were within range of findings from Navarrete et al. (Citation2016) (aucubin 1.78–3.80 g/kg DM and acteoside 0.5–41.7 g/kg DM). These compounds may help to explain the greater urine output seen for animals grazing plantain pastures. The concentration of aucubin and acteoside was greatest in late lactation, which coincided with higher urine outputs. Aucubin is known to stimulate the excretion of uric acid from the kidneys (Kato Citation1946). However, in the current experiment, the concentration of uric acid in the urine was lower for cows grazing pure plantain compared with those on pasture. It is unclear why we observed this result and further research may be necessary.
Despite greater total volume output, N output was not increased for cows grazing plantain. The lower urinary N concentration from cows grazing plantain was able to offset the increase in calculated or measured urine volume, resulting in a reduction in total N output when plantain was included in the diet. Using average urination size and assuming a patch size of 0.2 m2, the urine N loading from cows on perennial ryegrass-white clover pasture was approximately 700 kg N/ha in late lactation and 670 kg N/ha in early lactation. With the same assumptions, a urine patch from cows grazing plantain would have an N loading of approximately 450 kg N/ha in late lactation and 320 kg N/ha in early lactation. Application rates above 500 kg N/ha will likely increase leaching and nitrous oxide emission potential (Ledgard et al. Citation2007; Groenigen et al. Citation2010). This shows the potential of plantain to reduce N losses from grazing dairy systems. However, further research which defines what proportion of plantain in the diet is required to achieve reductions in N leaching and how to incorporate plantain into a grazing system is required.
Conclusions
Our results demonstrate a role for the use of plantain as a mitigation strategy to reduce the environmental impact of dairy farming. By providing plantain as a monoculture or with perennial ryegrass-white clover pastures to cows, milk solids production was increased or maintained and urine N concentration was reduced. The decline in urine N concentration may decrease urine patch N loading, thus reducing the risk of nitrate leaching for dairy grazing systems.
Acknowledgements
The programme is a partnership between DairyNZ, AgResearch, Plant & Feed Research, Lincoln University, Foundation for Arable Research and Landcare Research.
Disclosure statement
No potential conflict of interest was reported by the authors.
Additional information
Funding
References
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