Abstract [Objectives] The effects of metal elements in irrigation water in the tobacco areas of southern Anhui on the coke sweet aroma of tobacco leaves was determined.
　　[Methods] 53 representative areas for tobacco planting in southern Anhui were selected， and the quality of irrigation water， especially the content of metal elements， was investigated.
　　[Results] The contents of micro （medium） elements in the irrigation water were too low to have a significant effect on the formation of the coke sweet aroma style of tobacco leaves. The contents of Mg， Ca and Zn were 0.7-8.0， <40 and 0.002-0.029 mg/L， respectively. The heavy metal contents of the irrigation water and other basic control items all met corresponding national standards. Furthermore， the tobacco planting experiment under controlled irrigation using paddy soil in the greenhouse proved that Zn was a negative correlation factor for forming the coke sweet aroma style of tobacco and the threshold value was ≥10 mg/L in the irrigation water. Meanwhile， Mg was a positive correlation factor and the content of Mg to promote the coke sweet aroma style should be maintained at 40-90 mg/L. Ca and Mg had a synergistic effect， which was mainly appropriate for acid paddy soils.
　　[Conclusions] This study improves the quality and yield of the coke sweet aroma of tobacco leaves， and has important theoretical and practical value for the formation of a popular agronomic control method.
　　Key words Irrigation water; Metal elements; Burned sugar aroma style; Effect
　　
　　Received： August 12， 2020  Accepted： October 20， 2020
　　Supported by Anhui Southern Tobacco Co.， Ltd. （20150551004）.
　　Yakui CHEN （1990-）， female， P. R. China， engineer， devoted to research and development of contaminated soil remediation technology.
　　*Corresponding author. E-mail： [email protected]; [email protected].
　　 In 2003， flue-cured tobacco leaves with a coke sweet flavor characteristic similar to Zimbabwe tobacco leaves were found in the tobacco areas in southern Anhui[1]. After years of development， southern Anhui tobacco leaves have outstanding strong-flavor style and rich， delicate and distinctive characteristics of coke sweet aroma， and are well received by cigarette companies[2]. At present， the exploration of the reasons for the formation of the characteristic style of coke sweet aroma of tobacco leaves mainly focuses on soil characteristics[3]， meteorological factors[4]， ecological characteristics[2]， and as well as the effects of metal element content， carbon and nitrogen metabolism and aroma component content of tobacco leaves on the coke sweet aroma style[5-6]. Under the conditions of relatively consistent meteorological factors and climatic characteristics in the tobacco areas of southern Anhui， soil characteristics are the main reason for the coke sweet aroma of tobacco leaves， and the coke sweet aroma style of tobacco leaves is in order of sandy loam>paddy soil>silty sandy soil[7]. However， the total area of tobacco fields in southern Anhui is 462 700 hm2， of which paddy fields （tobacco-rice rotation） are 351 400 hm2， dry fields are 111 400 hm2， and sandy loam is distributed in a few areas. 　　In order to make better use of the unique and high-quality tobacco planting conditions in the southern Anhui tobacco areas and further improve the quality and yield of coke sweet tobacco leaves， creating an agronomic control method that can demonstrate the coke sweet aroma of tobacco grown in paddy soil has important theoretical and practical value. There is a close relationship between the contents of metal elements in soil and tobacco leaves[8]， which both have a certain degree of influence on the quality of flue-cured tobacco[9-10]. Under the premise that the soil texture cannot be changed on a large scale， it may be a feasible and controllable agronomic method to adjust the metal elements in irrigation water to highlight the coke sweet style of tobacco leaves. Firstly， we should study the effect of metal elements in irrigation water on the coke sweet style of tobacco leaves.
　　In this study， 53 representative tobacco planting areas in southern Anhui were selected to investigate the quality of irrigation water in each area， especially metal elements in the irrigation water， and the coke sweet aroma of tobacco leaves produced in each area was detected. Through the experiment of tobacco leaf planting with controlled irrigation water in the paddy soil in a greenhouse， the effects of metal elements in the irrigation water on the characteristic coke sweet style of tobacco leaves were explored， aiming to form a popular agronomic regulation method.
　　Materials and Methods
　　Experimental materials
　　After field investigations， the typical tobacco planting areas in Xuanzhou District， Langxi County， Guangde County， Jing County， Jingde County， Wuhu County， and Nanling County under the jurisdiction of the southern Anhui tobacco area were selected， and 53 fixed-point observation and sampling points were set up. According to the requirements of HJ25.2-2014 Site Technical Guidelines for Environmental Site Monitoring， water samples were collected in such three periods as before transplanting， vigorous growing period and tobacco leaf harvest， and the test results were averaged. Moreover， tobacco samples were collected from the sampling points for smoking.
　　The soil for the tobacco planting experiment with controlled irrigation water in a greenhouse was taken from the plough layer of the clay soil in the tobacco-rice rotation fields around the southern Anhui Tobacco Huwan Science and Technology Demonstration Park. The irrigation water was based on the groundwater from the demonstration park， which was added with ZnSO4·7H2O， CaSO4·2H2O and MgSO4·7H2O to make Zn2+， Ca2+ and Mg2+ in the irrigation water in the concentration ranges of 5-15， 300-900 and 40-140 mg/L， respectively. The tobacco leaf variety was Yunyan 97， which was the same as the tobacco fields surrounding the demonstration park. 　　Experimental methods
　　Testing of irrigation water quality
　　The contents of total cadmium （Cd）， lead （Pb）， total arsenic （As）， copper （Cu）， zinc （Zn）， and selenium （Se） of heavy metals in the water samples were tested by corresponding national standard methods， referring to GB/T 7475-1987， GB/T 7485-1987 and GB/T 11902-1989 for details. The contents of trace elements iron （Fe）， magnesium （Mg）， calcium （Ca）， and manganese （Mg） in the water samples were determined by inductively coupled plasma mass spectrometry （ICP-MS）. The water samples’ BOD5， COD， suspended solids， pH， total salt content， chlorides and other mandatory test items specified in the farmland irrigation water quality standards were tested using the corresponding national standard methods， referring to GB/T 7488-1987， GB/T 11914-1989， GB/T 11901-1989， GB/T 7494-1987， GB/T 13195-1991， GB/T 6920-1986， HJ/T 51-1999， GB/T 11896-1989 and GB/T 16489-1996 for details.
　　Sensory evaluation
　　Sample preparation was carried out according to the requirements of GB2635-1992 and Cigarette Technological Specifications[11]， and more than 4 experts were hired for cigarette sensory evaluation and scoring according to aroma quantity， aroma quality， concentration， strength， offensive odor， aftertaste， and coke sweet aroma style.
　　Tobacco planting experiment with controlled irrigation water in a greenhouse
　　The experiment was carried out in the nursery greenhouse of the southern Anhui Tobacco Huwan Science and Technology Demonstration Park. In order to prevent the mutual influence between the test plants， the plant spacing was increased to three times that of normal field planting. The planting time， fertilizing amount and management methods were the same as those of field tobacco planting. Because there was no rainwater supplement in the greenhouse， the water required for tobacco leaf growth （including setting root water） was all supplemented with artificially prepared irrigation water containing different micro （medium） elements by way of root irrigation. Each test site included 5 tobacco leaves， and the test was set with 3 parallel groups in random arrangement.
　　Results and Analysis
　　Investigation of irrigation water quality in the tobacco areas in southern Anhui
　　Heavy metal contents of irrigation water in southern Anhui
　　The heavy metals Cd， Pb， Cu， Zn and the rare earth elements As and Se are the basic controls stipulated in the national standard GB 5084-2005 Standards for Irrigation Water Quality， i.e.， the heavy metals and rare earth elements that must be detected for the safety judgment of the water quality of farmland irrigation water. The contents of Cd， Pb， and As in the irrigation water of the tobacco areas in southern Anhui were basically lower than the detection limits of the corresponding national standard detection method， indicating that the contents of Cd， Pb and As in the irrigation water of the tobacco area in southern Anhui were low. The contents of heavy metals Cu and Zn in the irrigation water samples taken from the 53 typical tobacco planting areas were， respectively， 0.001-0.005 and 0.002-0.029 mg/L， which were far lower than the irrigation water quality standards stipulated by national standards （1 and 2 mg/L， respectively）. The rare earth element Se is often monitored simultaneously with heavy metal elements in irrigation water due to its toxic effects on humans and crops at high concentrations. The Se content of irrigation water samples taken from the 53 typical tobacco planting areas was 0.001-0.003 mg/L， which was basically only 1/10 of the national standard value of 0.02 mg/L. 　　Micro （medium） element contents of irrigation water in the tobacco areas of southern Anhui
　　Mn， Ca， Mg， Fe， Cu and Zn are considered to be the micro （medium） elements necessary for tobacco leaf growth. Irrigation water is an important source for tobacco leaves to absorb these elements. Mn has important physiological functions in the growth， development and metabolism of tobacco plants. It is an activator of some synthase and synthetase in tobacco plants. It can participate in photosynthesis and chloroplast structure， promote carbon assimilation， and regulate some biochemical reactions， affecting the respiration of tobacco plants. It is generally believed that within the Ca concentration in irrigation water ranging from 75 to 900 mg/L， the yield of tobacco shows a parabolic change. When the mass concentration of Ca reaches 150 mg/L， the yield reaches the maximum， and when it exceeds 150 mg/L， it decreased instead. When Ca reaches 900 mg mg/L， the growth of tobacco plants is severely inhibited， and the tobacco leaves have poor taste， poor combustibility， and black ash color[12]. The test results showed that the Ca contents in the irrigation water samples of the 53 typical tobacco planting areas were generally less than 40 mg/L， which theoretically would not have a significant impact on tobacco growth and smoking characteristics. Zhu et al.[13] found that when the Mg2+ concentration in irrigation water was 136 mg/L， the dry matter accumulation， the maximum accumulation of nutrients and the maximum growth rate in flue-cured tobacco reached the optimal value， and the tobacco leaves could be fully burned， showing a good ash color. The test results showed that the Mg2+ content of the irrigation water samples in the tobacco area of southern Anhui ranged from 0.7 to 8.0 mg/L， which theoretically has a weak effect on the growth and smoking characteristics of tobacco leaves.
　　Test results of other basic control items of the irrigation water samples
　　As shown in Table 1， the detection results of BOD5， COD， suspended solids， pH， total salt， chloride and other basic control items in the irrigation water samples from the 53 typical tobacco planting areas of southern Anhui were all lower than the national standards specified in GB 5084-2005 Standards for Irrigation Water Quality， indicating that the ecology of the tobacco area in southern Anhui is well maintained， and the irrigation water fully meets the national water quality requirements for farmland irrigation water. Meanwhile， the test results showed that the total salt contents of the irrigation water samples from the 53 typical tobacco planting areas in southern Anhui were only 334-1 050 mg/L， proving that the irrigation water in the tobacco area of southern Anhui belongs to the typical "soft water" characteristics of southern China， and there was a problem that the contents of micro （medium） elements were generally low. 　　Correlation analysis between the contents of micro （medium） elements in irrigation water and the coke sweet aroma of tobacco leaves in southern Anhui
　　The contents of micro （medium） elements in the tobacco leaf samples from the 53 typical tobacco planting areas in southern Anhui
　　As shown in Table 2， the contents of micro （medium） elements in tobacco leaves produced in the tobacco area of southern Anhui were generally low， which might be directly related to the lower contents of micro （medium） elements in the irrigation water in the tobacco areas of southern Anhui. Among them， the contents of Cu， Fe， Mn and Zn were lower， which theoretically has little effect on the quality of tobacco leaves. The Zn content of tobacco leaves produced in the southern Anhui tobacco areas was slightly higher than the Zn content range of high-quality tobacco， which might have an adverse effect on the quality and the coke sweet characteristic of tobacco leaves， which should be taken seriously. The contents of Ca， K， and Mg elements in tobacco leaves produced in the southern Anhui tobacco areas were generally low. The difference was that the Ca content in most tobacco leaves was basically within the Ca content range of high-quality tobacco leaves， but the contents of K and Mg elements were significantly lower than the average levels of high-quality tobacco leaves in China.
　　 The Mg element participates in the formation of chlorophyll and is an essential element for the normal structure of chloroplasts. Its proportion accounts for about 2.7% of the molecular weight of chlorophyll[14]. Mg is also involved in the synthesis of carbohydrates， fats and lipids， proteins and nucleic acids[15]. Therefore， as one of the essential nutrients for flue-cured tobacco， Mg directly affects the yield and quality of tobacco leaves. Tobacco-growing soils in southern China are lacking in Mg[16]， while the tobacco planting process in the southern Anhui tobacco areas mainly focuses on the regulation of K and N fertilizers， and the Mg2+ content of the irrigation water samples in the southern Anhui tobacco areas was tested to be 0.7-8.0 mg/L， at a relatively low level. It is inferred from this that the regulation of the Mg element in the irrigation water might be one of the research directions to highlight the coke sweet aroma of tobacco leaves.
　　Correlation between the contents of micro （medium） elements in the irrigation water and the coke sweet aroma of tobacco leaves in the tobacco areas of southern Anhui 　　After the tobacco leaves produced in the 53 typical tobacco planting areas were harvested and roasted， 500 g of the representative central tobacco leaves of grade C3F was selected from each area. After sample preparation， 4 or more experts smoked and evaluated the tobacco. The arithmetic average of the expert smoking results was used as the final score of the tobacco leaf samples. Among them， the tobacco leaf samples with a coke sweet aroma greater than 5 points are shown in Table 3.
　　The tobacco leaves produced in the 53 typical tobacco planting areas in southern Anhui selected were all identified as the coke sweet aroma type， and 8 of the tobacco leaf samples were scored greater than 5 points for the coke sweet aroma， which belonged to the more prominent coke sweet aroma type. The eight tobacco leaf samples were mainly distributed in Xuanzhou District and Nanling County. Geographical location and climatic characteristics might be the main factors affecting the quality of the coke sweet aroma. Among the eight tobacco leaf samples， the number of tobacco leaf samples produced from sandy loam soil accounted for the majority. Soil characteristics might be another important reason for the quality of the coke sweet aroma. Regardless of whether the tobacco leaves were produced from sandy loam soil or clay soil， the corresponding Mg content of tobacco leaves and irrigation water varied greatly. The reason might be that the contents of micro （medium） elements in the irrigation water and tobacco leaf samples in southern Anhui were generally low， which did not reach the concentration ranges that have a significant effect on the coke sweet smell of tobacco leaves. The coke sweet aroma of tobacco leaf samples might be the result of a combination of many factors， among which the micro （medium） element contents of the irrigation water played a limited role. However， the adjustable ranges of ecological factors， genetic factors and cultivation factors of tobacco planting are limited， and it is difficult to form adjustable agronomic methods.
　　Tobacco planting experiment using controllable irrigation water
　　In order to avoid the influences of the ecological factors， soil characteristics and other cultivation factors of tobacco planting and explore the possibility of adjusting the micro （medium） elements of irrigation water to enhance the coke sweet aroma of tobacco leaves planted in the paddy soil， a specific paddy soil was selected to carry out a tobacco planting experiment in a greenhouse using the prepared irrigation water containing different micro （medium） elements， and then the effects of the contents of micro （medium） elements in the irrigation water on the coke sweet aroma of tobacco leaves were analyzed. The basic properties of the paddy soil used in the experiment were： soil texture （clay）， organic matter （34.5 g/kg）， alkali-hydrolyzable nitrogen （201.4 mg/kg）， available phosphorus （35.71 mg/kg）， and available potassium （278.3 mg/kg）， and pH value 5.5， and the background values of micro （medium） elements in the groundwater （source of irrigation water） were： iron （0.012 mg/L）， copper （0.007 mg/L）， zinc （0.010 mg/L）， manganese （0.065 mg/L）， calcium （15.46 mg/L）， magnesium （3.625 mg/L）， and pH 6.7. The smoking results of the tobacco leaf samples produced are shown in Table 4. 　　As shown in Table 4， when the concentration of Zn2+ in the irrigation water was greater than 10 mg/L， it had an adverse effect on the coke sweet aroma of the tobacco leaves. When the concentration of Zn2+ in the irrigation water was 15 mg/L， the coke sweet aroma of the tobacco leaves dropped from 4.6 to 4.1 points. GB 5084-2005 Standards for Irrigation Water Quality stipulates that the Zn2+ concentration in irrigation water is ≤2 mg/L， and the Zn content in the irrigation water in the tobacco area of southern Anhui obtained from the investigation is much lower than this value. When the concentration of Mg2+ in the irrigation water was 40 and 90 mg/L， the core sweet aroma of tobacco leaves increased from 4.6 to 4.8 and 5.0 points， respectively， indicating that Mg was a positively correlated factor for the coke sweet flavor of tobacco leaves.    As mentioned earlier， Mg might participate in the formation of chlorophyll in tobacco leaves， thereby affecting the accumulation of total sugar in tobacco leaves and increasing the intensity of sweetness. The investigation results showed that the Mg2+ content of the irrigation water in the tobacco area of southern Anhui ranged from 0.7 to 8.0 mg/L， much lower than the experimental value. In order to show the core sweet smell of tobacco leaves in southern Anhui， the Mg element should be appropriately supplemented in the irrigation water. When the Mg2+ concentration in the irrigation water was 140 mg/L， the coke sweet aroma of the tobacco leaves began to show signs of decline， which might be related to the increase in the overall salt concentration. Therefore， the content of Mg element in irrigation water should be maintained at 40-90 mg/L.
　　 When the concentration of Ca2+ in the irrigation water was 300 and 600 mg/L， the coke sweet aroma of tobacco leaves was improved to a certain extent， but when the concentration of Ca2+ in the irrigation water was 900 mg/L， the quality of the coke sweet aroma of tobacco leaves was significantly reduced， from 4.6 points in the control to 4.0 points. Ca element is the main component of the ash of tobacco leaves after burning， and it is generally believed that Ca element does not participate in the composition of tobacco leaves and affect the formation of other functional groups or organelles of tobacco leaf cells. When the content of Ca in the irrigation water is high， the growth of tobacco plants is severely inhibited， and the tobacco leaves have poor taste， poor combustibility， and dark ash. The tobacco-growing soils in the tobacco areas of southern Anhui are mainly clay soils for tobacco-rice rotation， which have high viscosity， high soil organic matter content， and less air permeability than sandy loam， and the overall pH is acidic. Appropriate increase of Ca2+ content in the irrigation water helps increase soil pH. In addition， the combination of Ca2+ and soil organic matter can increase the particle size and air permeability of the soil， which is conducive to the formation of the core sweet smell of tobacco leaves. 　　Meanwhile， the contents of Mg and Ca elements in the irrigation water could be increased. For example， when the concentration of Mg2+ was 90 mg/L and the concentration of Ca2+ was 600 mg/L， the coke sweet aroma of tobacco leaves increased from 4.6 to 5.3 points， indicating that Mg and Ca had a synergistic effect on the coke sweet aroma of tobacco leaves. It is generally believed that Mg2+ and Ca2+ are elements of the same family， and tobacco leaves have a competitive effect on the absorption of Mg2+ and Ca2+. The synergistic effect obtained in this study might be due to the overall low contents of Mg and Ca in the soil and irrigation water in the tobacco areas of southern Anhui.
　　Conclusions and Discussion
　　The contents of micro （medium） elements in the irrigation water from the 53 representative tobacco planting areas in southern Anhui were generally low， which is consistent with the basic characteristics of "soft water" in southern China. For example， the contents of Mg element in the irrigation water in the southern Anhui tobacco areas ranged from 0.7 to 8.0 mg/L， the contents of Ca were generally less than 40 mg/L， and the contents of Cu and Zn were 0.001-0.005 and 0.002-0.029 mg/L， respectively.
　　The contents of heavy metals and basic control items such as BOD5， COD， suspended solids， pH， total salt content and chloride in the irrigation water samples of the tobacco areas in southern Anhui were all lower than the national standards specified in GB 5084-2005 Standards for Irrigation Water Quality， indicating that the ecology of the tobacco area in southern Anhui is maintained well， and the safety of irrigation water fully meets national standards.
　　There was no obvious correlation between the contents of micro （medium） elements in the irrigation water from the 53 representative tobacco planting areas in southern Anhui and the coke sweet aroma of the tobacco leaves. The contents of micro （medium） elements were generally low and did not reach the concentration ranges that have a significant effect on the coke sweet aroma of tobacco leaves.
　　Through the tobacco planting experiment in the greenhouse using paddy soil， the results showed that Zn in the irrigation water was a negatively correlated factor of the coke sweet aroma of tobacco leaves， and the threshold of impact was 10 mg/L. Mg was a positively correlated factor， and the content of Mg element in the irrigation water that highlights the coke sweet aroma of tobacco leaves should be maintained at 40-90 mg/L. Ca and Mg had a certain synergistic effect， which is mainly suitable for acidic compact clay such as paddy soil. 　　References
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　　Editor： Yingzhi GUANG  Proofreader： Xinxiu ZHU