Agricultural and Food Science in Finland, Vol. 12 (2003): 107–115 107 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Vol. 12 (2003): 107–115. Effect of liming on yield and quality of peppermint and Sachalin mint in fine sand soil of Northern Finland Abbas Aflatuni MTT Agrifood Research Finland, North Ostrobothnia Research Station, FIN-92400 Ruukki, Finland, e-mail: abbas.aflatuni@mtt.fi Jouko Uusitalo, Sari Ek University of Oulu, Laboratory of Mass Spectrometry, Department of Chemistry, FIN-90014 Oulu, Finland Anja Hohtola University of Oulu, Department of Biology, Botany, FIN-90014 Oulu, Finland Soil acidity commonly limits plant production in the fine sand soil of Northern Finland, which often has a low pH (5.5–6.5) and contains low levels of Ca and Mg. The effect of five liming (10% Mg and 19% Ca) levels, 0, 4, 8, 12, and 16 tons ha-1, on the herb and essential oil yield and menthol and menthone content of two mint species (peppermint, Mentha x piperita, a variety of Black Mitcham and Sachalin mint, Mentha arvensis var. sacchalinensis) cultivated in fine sand soil in Northern Fin- land (64°40’N and 25°05’E) was studied during 1998–2000. Liming clearly increased the pH levels and the Ca and Mg content of the soil. The dry matter content, essential oil quantity, and the menthol or menthone content in mints were not affected by liming. In comparison with no liming however, liming at a rate of 4 t ha-1 doubled the herb yield. The highest yield was achieved in Sachalin mint by liming at 4 or 8 t ha-1 in the second and third year (soil pH 6–6.5) (Ca 725–871 mg l-1 and Mg 122– 219 mg l-1), and in peppermint by liming at 4, 8 or 16 t ha-1 (soil pH 6–6.6) (Ca 725–1272 mg l-1 and Mg 122–245 mg l-1). Therefore, we conclude that a higher peppermint and Sachalin mint yield is achieved by increasing soil pH to values above 6.0 in the fine sand soil of Northern Finland. Key words: Lamiaceae, lime, Mentha arvensis var. sacchalinensis, Mentha x piperita, menthone, menthol, pH, roots, stolons © Agricultural and Food Science in Finland Manuscript received October 2001 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Vol. 12 (2003): 107–115. Introduction Over the past few years, different herb species have been a subject of continuous interest to con- sumers, growers, and researchers in Finland. Peppermint (Mentha piperita L.), a member of the Lamiaceae family, is a rhizomatous aromat- ic plant widely cultivated in many countries, such as the USA, India, China, the former USSR, Ita- ly, France and Hungary. Its essential oil is con- sidered industrially important (Lawrence 1985) mailto:abbas.aflatuni@mtt.fi 108 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Aflatuni, A. et al. Effect of liming on two mint species as it is used in pharmaceuticals, confectionery, alcoholic beverages, chewing gum, dental creams and cosmetics, and its leaves are used in food flavoring, tea and folk medicine (Gupta 1991). Each year, Finland imports about 8 t of dry mint leaves, 10–20 t of peppermint essential oil, and 10 t of pure menthol (National Board of Cus- toms 1984–1996). Many reports on the benefits of long days of sunlight for mint cultivation (Bur- bott and Loomis 1967, Clark and Menary 1980) make increasing Finnish production an attrac- tive proposition. Soil pH profoundly affects the growth and nutrient uptake by crops. Shukla et al. (1997, 1998) observed that different levels of soil pH influenced the fresh and dry weight and the es- sential oil yield of peppermint. It has been found that high levels of Mg in nutrient solu- tion increase the leaf essential oil content of Mentha arvensis without significantly altering the quality of the essential oil (Zimma and Pie- kos 1988, Maia et al. 2001). The leaves of pep- permint are rich in K and Mg (Berbec and Kolodziej 1996). One of the factors limiting plant production in Finland is soil acidity; the average pH value of agricultural soils in 1981–1987 was 5.8 (Kähäri et al. 1987). The pH range in the fine sand soil of Northern Finland is 5.5–6.5, and lim- ing is commonly needed. The acidic sandy soils of Northern Finland are also poor in Ca and Mg. It is to be expected that liming the soil is neces- sary in order to profitably cultivate mints in acid- ic soils. At present, the cultivation of mint is very lim- ited in Finland. In order to offer farmers in North- ern Finland a new source of income, a project was started to examine the optimal conditions for the cultivation of mint. Because of favorable light conditions (long days) and cool nights, high quality essential oil yield can be expected (Franz 1984). In this particular study, the impact of lim- ing (including the manipulation of soil pH and the contents of Ca and Mg) on the yield and qual- ity of Mentha x piperita and Mentha arvensis var. sacchalinensis species was investigated dur- ing a three-year field experiment. Material and methods Plant material and the field experiment The experiment was established in 1998 in Ruuk- ki (Northern Finland) at the North Ostrobothnia Research Station of MTT Agrifood Research Finland, 64°40’N and 25°05’E. It was set up in a randomized split-plot design of four blocks (Gomez and Gomez 1984), where the main plots were laid out in each block according to a rand- omized complete block design. There were two main plots (mint species) in each block: 1) Men- tha x piperita “Peppermint” (a variety of Black Mitcham, originally from Egypt) and 2) Mentha arvensis var. sacchalinensis (originally from Hungary). Table 1 shows the experimental field treat- ment dates in 1998–2000. The amount of N, P, and K applied annually were 40, 28, and 56 kg ha-1, respectively. The lime was spread manual- Table 1. The treatment dates of the field experiments in 1998–2000. Treatment 1998 1999 2000 Soil sampling 28 May and 15 Sep 11 May Fertilizer application 7 Jun 7 Jun 22 May Liming 7 Jun – – Planting 9 Jun – – Harvesting 27 Aug 17 Aug 15 Aug Root and stolon development observation 16 Aug 109 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Vol. 12 (2003): 107–115. ly before fertilization and mixed immediately after being spread with an S-spike. The subplots received 0, 4, 8, 12, or 16 t ha-1 of lime. The lime was provided by Saxo Mineral Oy in Tornio (Northern Finland). The lime contained more than 10% Mg and more than 19% Ca. The fast acting neutralizing capability of liming was 35% of the material, according to the Finnish pH Stat Method. Determination of the neutralizing val- ue was performed by the Finnish Plant Produc- tion Inspection Center (unpublished, as described by Erstad et al. 2000). The plants were micropropagated. Nodal cut- tings were cultured on Murashige and Skoog’s (1962) medium for shoot regeneration and on Lloyd & McCown’s woody plant medium (1980) for rooting. The rooted plants were planted after fertilization and liming in the experimental field in spring 1998. The plot size was 1.5 x 5 m (= 7.5 m2); the distance between the rows was 75 cm and the distance between the plants in the rows was 20 cm. Analyses Samples for soil analyses were taken in May 1998 before liming and fertilization and in Sep- tember 1998 and May 2000 after liming and fer- tilization (Tables 1 and 2). Five subsamples were taken from each subplot replication (liming lev- el) and mixed in order to obtain one sample of approximately 0.5 l from each liming level. The sampling depth was 0–20 cm. In this experiment, an analysis of statistical error was not possible because only one sample per liming level was taken. The samples were analyzed by Soil Analysis Service Ltd. The soil texture and organic matter content were estimated by finger assessment. The soil texture was that of fine sand containing 6– 12% of organic matter. To analyze soil nutrients, 25 ml of soil was extracted with 250 ml of acid- ic ammonium acetate (pH 4.65) (Vuorinen and Mäkitie 1955) and a plasma emission spectrom- eter was used for the determinations. A spectro- photometer was used for the determination of P. After standing 12 hours, the soil pH and electri- cal conductivity (EC) were determined in soil- water suspension (1:2.5). A specific electrode was used to measure both the pH and EC in this suspension.The results of the analyses before liming and fertilizing were: soil texture, fine sand with 6–12% of organic matter content and elec- trical conductivity, 0.8 mS cm-1, pH 5.8, and Ca 677, P 21, K 214 and Mg 41.6 mg l-1 of soil. The fresh yield was determined after harvest: dried (at +40°C) random samples (2 x 200 g) were used to assess dry matter content and to estimate the concentration of essential oil. To Table 2. The results of soil analyses after liming and fertilizing in 1998 and 2000. Analysis Liming level t ha-1 0 4 8 12 16 September 1998 pH 5.5 6.0 6.1 6.4 6.5 Ca mg l-1 569 725 836 979 1270 P mg l-1 26 25 25 25 26 K mg l-1 141 119 128 127 138 Mg mg l-1 48 122 168 202 223 May 2000 pH 5.5 6.0 6.5 6.6 6.6 Ca mg l-1 527 754 871 892 1272 P mg l-1 23 25 24 22 24 K mg 1-1 197 199 203 194 192 Mg mg l-1 72 176 219 214 245 110 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Aflatuni, A. et al. Effect of liming on two mint species isolate the essential oil, mint leaves (50 g) were dried at +30°C and then mixed with 700 ml of water which was subsequently hydrodistilled for 2 h at 120°C at atmospheric pressure. The es- sential oil content was then measured (Pohjamo 1994). The constituents of the essential oil were an- alyzed using GC and identified by GC/Ms. Sam- ple preparation: Leaf samples, which were se- lected to represent leaves of all ages, were col- lected at random from randomly selected plants. Gas chromatography was performed with a Per- kin Elmer Autosystem XL gas chromatograph using helium as the carrier gas at a constant pres- sure mode (37 psig). The sample components were identified by mass spectra matching with a Wiley/NBS Reg- istry Mass Spectra Data Base. The identifications were verified by comparing the retention time and mass spectrum with, when possible, a refer- ence compound. Root and stolon observation When finishing the experiment, root and stolon samples were taken from each plot in the year 2000 in order to observe their development. The root density, the number of stolons, as well as the weight of roots and stolons, were measured. The density of roots and the number of stolons were evaluated on a scale of 1–4, in which 1 was remarkably few, 2 few, 3 optimum and 4 many. The observation dates are shown in Table 1. Weather conditions during the experiment The monthly average temperature in the summer 1998 was lower than normal in North Ostroboth- nia and in the summer 1999, it was close to nor- mal; in the year 2000, May and July were higher than normal (Table 3). The precipitation in the summers of 1998 and 2000 were higher than normal and in 1999, it was close to average. Statistical methods The randomization method led to the split-plot experimental design. Hence, the response varia- bles were analyzed using the traditional ANO- VA for split-plot design (Gomez and Gomez 1984). Measurements of fresh yield and essential oil content were repeated several times for each plot. The repeated measurements tended to correlate, which was taken into account in the model used. The covariance structure of the repeated meas- urements was chosen by comparing potential structures using Akaike and Schwarz’s Bayesian information criterion (Wolfinger 1996). The model, and the assumptions used, was illustrat- ed by Gumperetz and Brow (1993). T h e a s s u m p t i o n s i n b o t h m o d e l s w e r e checked graphically by box-plot in order to test the normality of errors; plots of residuals were used to test the constancy of error variance (Neter et al. 1996). The parameters of the models were estimated by the restricted maximum likelihood (REML) estimation method using the SAS sys- tem for Windows, release 8.1. Results and discussion Impact of liming on soil Liming clearly increased the Ca and Mg content of the soil (Table 2). In Finland, there are no rec- Table 3. The monthly mean summer temperatures (°C) in 1998–2000 and long-term average in Ruukki. Data provid- ed by the Finnish Meteorological Institute. Year Month 1998 1999 2000 1961–90 May 6.3 5.4 8.9 7.7 June 12.7 15.5 13.1 13.2 July 15.4 15.8 16.3 15.4 August 12.4 11.7 13.1 13.1 111 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Vol. 12 (2003): 107–115. ommended soil nutrient levels for herbs. Accord- ing to Soil Analysis Service Ltd (1997), a Ca content of 1400–2000 mg l-1 and an Mg content of 120–200 mg l-1, which is the targeted level, are satisfactory in fine sand soil used for grow- ing outdoor vegetables. The Ca and Mg levels in the experimental field were initially lower than the ranges recommended by Soil Analysis Serv- ice (see Table 2) but the satisfactory values were achieved after liming. In 1998, the soil pH in plots after liming with 4, 8, 12, and 16 t ha-1 were respectively 0.5, 0.6, 0.9, and 1.0 units higher than in plots with no liming: the pH range in liming levels in 1998– 2000 was 6–6.6. Although liming was performed only in 1998, the soil pH levels in plots limed with higher amounts than 4 t ha-1 seemed to be higher in 2000 than they were in 1998, probably due to the slow dissolution of lime. Similarly, according to Kemppainen et al. (1993), after lim- ing with 8 or 12 t ha-1 in the first year of trials, the pH level increased during the 7 years of ex- periment in fine sand while at 4 t ha-1 pH level, the pH rose only in the first year and remained the same during the 7 years. The recommended pH value for all soils is 6.5–7 (Soil Analysis Service 1997). Yield, oil content, and menthol and menthone content Liming had the most pronounced effect on fresh yield (Table 4). In 1999, the fresh yield of Sacha- lin mint was significantly, and for peppermint almost significantly, higher with liming in com- parison with no liming (Table 4). In 2000, the yields were much lower than in the two previous years, which was due to weeds, bad overwintering, and plant age. Galambosi (1995) also found a decrease in yields of bio- logical mint in trials performed in South Mikke- li, Finland. Moreover, according to Hornok (1992), a peppermint plantation should be main- tained for two, or rarely, for three years and it should not be cultivated in the same field for four years due to increases of weeds, insects, and dis- eases. In our experiment, however, all plants were healthy and no signs of disease were ob- served. In this experiment, the effect of liming on the fresh yield of Sachalin mint was almost signifi- cant. The yield of Sachalin mint was higher with liming at a level of 4 or 8 t ha-1 (Soil pH 6–6.5) in comparison with no liming (soil pH 5.5). The highest yield of peppermint was also achieved Table 4. Fresh yield of the Sachalin mint and peppermint (kg ha-1) in 1998–2000 at different liming levels. Species Liming level t ha-1 P-value* 0 4 8 12 16 Year 1998 Sachalin mint 7867 12267 9600 13334 12400 0.06 Peppermint 16267 16667 16933 18266 17200 0.88 Mean 12000 14400 13333 15733 14800 0.11 Year 1999 Sachalin mint 10933 24800 23733 23333 27866 <0.005 Peppermint 9200 16333 15867 14800 17600 0.05 Mean 10133 20400 19733 19067 22666 <0.001 Year 2000 Sachalin mint 2800 7200 6600 2533 2133 0.05 Peppermit 1333 2120 2016 1466 1867 0.06 Mean 2060 4550 4250 1985 1888 0.08 P-value* = P-value between five liming level differences 112 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Aflatuni, A. et al. Effect of liming on two mint species with the liming level of 4 or 8 in comparison with no liming, but only in the second and third years (1998 and 2000) after application. The average yield for three years shows that liming with 4 t ha-1 gave almost the same yield as with 8 and 12 t ha-1. Matusiewicz (1972) studied pep- permint of the Mitcham variety for three years in five combinations of soil pH, ranging from 4.7 to 6.9. He concluded that the plants devel- oped best and gave the highest crop of plant material at soil pH ranging from 5.6 to 6.2. In addition to low pH, the reason for low yield could also be Ca and Mg deficiency, which limited the yield in treatments when liming was not applied. The dry matter content for Sachalin mint in 1998, 1999, and 2000 was 17–18%, 18–19%, and 17–19%, and for peppermint it was 13–14%, 19– 20%, and 19–21%, respectively. Liming did not have any effect on the percentage of dry matter. Differences in dry matter were seen between dif- ferent species (P < 0.0001) and between differ- ent years (P < 0.0001). Liming did not affect the essential oil con- tent of Sachalin mint (P = 0.38 in 1998 and P = 0.16 in 1999,) and peppermint (P = 0.16 in 1989 and P = 0.71 in 1999,) (Fig. 1). The slightly (0.02) higher essential oil yield was achieved in 1998 by liming in Sachalin mint (Fig. 2). In 1999, although the essential oil yield was higher with liming than without it, the differences in both mints were not significant because of the wide random variation. Wide random variation in es- sential oil yield is due to factors in the field oth- er than genetic differences, as the plant material used in this experiment was micropropagated. The menthol and menthone content of Sachalin mint and peppermint were not influenced by lim- ing (Table 5). Development of rootage Both the deficiency and excess of minerals has a negative effect on root growth. The effect of liming on the weight and density of roots in the soil was not significant, but Sachalin mint roots were longer (P < 0.001) in limed plots at the pH 6–6.5, in comparison with unlimed plots (pH 5.5). In peppermint, the difference in root length was not statistically significant (Table 5). Fig. 1. Essential oil content (%) in dry leaves of Sachalin mint and peppermint in 1998–1999 at dif- ferent liming levels. 113 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Vol. 12 (2003): 107–115. Fig. 2. Essential oil yield (kg ha-1) of Sachalin mint and peppermint in 1998–1999 at different liming levels. Table 5. The growth of mint rootage and contents of menthol and menthone (%) of total peak area at different liming levels. Liming levels t ha-1 Significance 0 4 8 12 16 SEM P1 Sachalin mint Root density per plant, 1–4 scale* 1.8 2.0 2.3 1.8 2.0 0.28 0.25 Number of stolons per plant, 1–4 scale* 1.5 2.3 2.0 1.8 2.0 0.29 0.14 Root and stolons, g per plant 124 162 277 135 235 79.0 0.38 Root length, cm per plant 11.9 15.8 18.3 16.5 22.8 2.44 <0.001 Menthol % 82.5 81.8 80.8 82.5 82.9 1.53 0.88 Menthone % 10.5 8.90 9.20 8.40 7.60 1.09 0.54 Peppermint Root density per plant, 1–4 scale* 3.8 3.5 3.5 4.0 3.0 0.28 0.41 Number of stolons per plant, 1–4 scale* 3.5 3.0 3.8 4.0 3.0 0.29 0.85 Root and stolons, g per plant 656 642 599 736 441 79.0 0.56 Root length, cm per plant 19.5 22.3 21.5 22.8 20.3 2.44 0.40 Menthol % 53.5 46.3 51.3 48.9 52.0 3.94 0.72 Menthone 20.5 24.8 22.1 22.2 22.2 4.72 0.98 SEM = standard error of mean P1 = P value between 0 and other lime level Scale* 1 = remarkably few, 2 = few, 3 = optimum and 4 = abundant 114 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Aflatuni, A. et al. Effect of liming on two mint species Summary and conclusions By applying 4 t ha-1 of lime, the pH rose from 5.5 to 6 in fine sand soil with 6–12% organic matter content. Liming almost doubled the fresh herb yield of both Sachalin mint and peppermint. The essential oil yield was higher in Sachalin mint in the first year. Liming had no effect on the essential oil content and proportion of men- thol and menthone; neither did it have any ef- fect on the development of peppermint rootage. However, liming at the level of 4 or 8 t ha-1 (pH 6–6.l) did have a positive effect on the length of roots in Sachalin mint. In conclusion, if the soil pH value is lower than 6 or the Mg and Ca are low, liming at the level of 4–8 t ha-1 in the sandy soil of Northern Finland is recommended in order to achieve a higher fresh yield and a higher essential oil yield. Liming with 12 t ha-1 produces almost the same yield as with 4 t ha-1 during the first three years after liming. Liming has no effect on the essen- tial oil content, the proportion of menthol and menthone, or the percentage of dry matter. The roots of Sachalin mint are longer with liming. References Berbec, S. & Kolodziej, B. 1996. Macro- and microele- ment content in raw materials of some medicinal plants depending on the period of harvest. Annales Universitatis Mariae Curie Sklodowska 51: 55–61. Burbott, A.J. & Loomis W.D. 1967. Effects of light and temperature on the monoterpenes of peppermint. Plant Physiology 42: 20–28 Clark, R.J. & Menary, R.C. 1980. Environmental effects on pepper mint (Mentha piperita L.). I. Effect of daylength, photon flux density, night temperature and day temperature on the yield and composition of pep- permint oil. II. Effects of temperature on photosyn- thesis, photorespiration and dark respiration in pep- permint with reference to oil composition. Australian Journal of Plant Physiology 7: 685–697. Erstad, K.J., Konovalov, Y.N., Putro, J., Rex, M. & Luuk- konen, E. 2000. Reactivity of silicate liming materi- als from Northern Europe assessed by soil incuba- tion and two pH stat methods. Agricultural and Food Science in Finland 9: 333–348. Franz, C. 1984. Influence of the growing site on the quality of Mentha piperita L. oil used for pharmaceutical, cosmetic and flavoring. International Society for Hor- ticultural Science 144: 145–150. Galambosi, B. 1995. The organic cultivation of spices and medicinal herbs. Mauste- ja rohdosyrttien luonnon- mukainen viljely. Helsinki, Painatuskeskus, 234 p. ISBN 951-1530-2. Gomez, K.A. & Gomez, A.A. 1984. Statistical procedures for agricultural research. 2nd edition. John Wiley & Sons, New York. 627 p. Gumperetz, M.L. & Brow, C. 1993. Repeated measures in randomized block and split-plot experiments. Ca- nadian Journal of Forest Research 23: 625–639. Gupta, R. 1991. Agrotechnology of medicinal plants. In: Wijesekera, R.O.B. (ed.). 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Journal of Agri- cultural, Biological, and Environmental Statistics 2: 205–230. Zimma, D. & Piekos, R. 1988. Extraction of eight essen- tial elements from the leaves of peppermint, Mentha piperita (L.) Huds. Herba Hungarica 27: 65–75. SELOSTUS Kalkituksen vaikutus piparmintun ja Sachalinin mintun satoon Pohjois-Suomessa Abbas Aflatuni, Jouko Uusitalo, Sari Ek ja Anja Hohtola MTT (Maa- ja elintarviketalouden tutkimuskeskus) ja Oulun yliopisto Maan happamuus rajoittaa yleisesti kasvien tuotan- toa Pohjois-Suomen hienohietamailla, joiden pH vaihtelee 5,5–6,5 ja kalsium- ja magnesiumtaso ovat yleensä alhaisia. Kalkituksen vaikutusta kahden min- tun (piparminttu, Mentha x piperita, lajike Black Mitcham ja Sachalinin minttu, Mentha arvensis var. sacchalinensis) satoon sekä haihtuvan öljyn määrään, mentoli- ja mentonipitoisuteen tutkittiin Pohjois-Suo- messa vuosina 1998–2000. Kalkkia (10 % Mg ja 19 % Ca) levitettiin joko 0, 4, 8, 12 tai 16 t/ha. Maa- laji oli hienoa hietaa. Kalkitus nosti selvästi maan pH:ta sekä kalsium- ja magnesiumtasoa. Kalkitus ei vaikuttanut sadon kui- va-ainepitoisuuteen, öljyn määrään eikä mentolin tai mentonin pitoisuuteen. Neljä t/ha kalkkia saaneilta aloilta tuoresato oli kaksi kertaa suurempi kuin aloil- ta, joita ei oltu kalkittu. Sachalinin mintun sato oli suurin, kun sitä kalkittiin 4–8 t/ha toisena ja kolman- tena vuonna (maan pH 6–6,5, Ca 725–871 mg/l and Mg 122–219 mg/l). Piparmintun sato oli suurin, kun sitä kalkittiin 4, 8 tai 16 t/ha (maan pH 6–6,6, Ca 725–1272 mg/l ja Mg 122–245 mg/l). Näin ollen Pohjois-Suomen hienolla hietamaalla voidaan saada hyvä piparmintun ja Sachalinin mintun sato, kun maan pH on yli 6. 116 A G R I C U L T U R A L A N D F O O D S C I E N C E I N F I N L A N D Aflatuni, A. et al. Effect of liming on two mint species Title Introduction Material and methods Results and discussion Summary and conclusions References SELOSTUS