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托福阅读真题第7篇RegulationofBodyTemperature大多数动物都有可以生存的体温上限。
例如，在人类中，41摄氏度的体温会导致蛋白质功能丧失和神经系统崩溃，而42至43摄氏度的体温会导致是致命的。
鸟类的静息体温略高于哺乳动物（大约为40 至 41 摄氏度，而大多数哺乳动物为 35 至 38 摄氏度），它们无法在高于 46 至 47 摄氏度的体温下生存。
在高于 50 摄氏度的环境温度下，几乎所有动物都会死亡。
然而，在温度范围的另一端，极冷的耐受性更好。
例如，一些动物在解冻后可以冷冻并存活。
冷冻/解冻通常是危险的，因为冰晶会在细胞和破裂膜内形成。
然而，许多昆虫，如毛毛虫，少数种类的两栖动物，如林蛙，以及极少数爬行动物，如彩龟，都可以阻止其细胞中的晶体形成。
他们通过从肝脏中大量流出葡萄糖来响应皮肤表面的冰块来做到这一点。
储存在肝脏中的葡萄糖进入血液和细胞，降低它们的冰点，使细胞不会冻结成固体。
这些动物可以长时间完全冷冻 65% 或更多的身体，只有在温暖时期才解冻而不会产生有害影响。
他们通过从肝脏中大量流出葡萄糖来响应皮肤表面的冰块来做到这一点。
储存在肝脏中的葡萄糖进入血液和细胞，降低它们的冰点，使细胞不会冻结成固体。
这些动物可以长时间完全冷冻65% 或更多的身体，只有在温暖时期才解冻而不会产生有害影响。
他们通过从肝脏中大量流出葡萄糖来响应皮肤表面的冰块来做到这一点。
储存在肝脏中的葡萄糖进入血液和细胞，降低它们的冰点，使细胞不会冻结成固体。
这些动物可以长时间完全冷冻 65% 或更多的身体，只有在温暖时期才解冻而不会产生有害影响。
过去，动物被分为两类。
冷血动物需要外部热源（例如阳光）来温暖自己。
相比之下，温血动物使用内部热量来维持体温。
然而，这些术语具有误导性，因为许多冷血动物可以通过锻炼骨骼肌产生大量热量。
事实上，许多人在白天的体温至少与鸟类和哺乳动物等温血动物的体温一样温暖。
生物学家现在根据动物的热源和维持体温的能力对动物进行分类。
Impact of temperature on chemicalreactions随着气候变化的加剧，温度变化对化学反应的影响日益突显。
在这篇文章中，将探讨温度变化对化学反应的影响、温度与化学反应动力学参数之间的关系以及有关温度的实验设计和数据分析方法。
最后，将讨论如何利用这些信息来实现更有效的化学反应过程和更可持续的生产方式。
1. 温度对化学反应的影响温度是化学反应速率的最重要的因素之一。
热力学上，温度的提高会使反应物的反应自由能降低。
这意味着可以在更低的反应物浓度下实现更高的反应速率和更高的反应产率。
在实际应用中，这意味着可以使用更小的催化剂剂量、更低的反应温度和更短的反应时间来实现更高的化学反应效率。
2. 温度与化学反应动力学参数之间的关系根据阿累尼乌斯方程，反应速率常数k与温度T之间呈指数关系。
因此，反应速率常数随温度的升高而迅速增加。
然而，这并不意味着温度越高，反应速率越大。
随着温度继续上升，分子之间的碰撞会变得越来越频繁，而且分子的热能越来越高，从而导致分子分解并丢失它们可能已经获得的活性。
这种反应损失称为热分解。
因此，存在一个适当的反应温度，可以在其中实现最大的反应速率和最大的反应产率。
3. 有关温度的实验设计和数据分析方法在研究温度对化学反应的影响时，需要设计实验来控制反应温度，并记录反应速率和反应产物的浓度。
为了确定最佳的反应温度，可以使用Arrhenius方程来计算反应速率常数，并绘制不同温度下的反应速率常数随温度变化的图表。
这些数据可以用来计算反应的激活能，并使用线性回归分析来确定此数据的拟合度和预测温度下的反应速率常数。
4. 应用温度控制化学反应化学反应的有效控制和优化对于实现可持续生产和生产高质量产品至关重要。
利用温度控制策略可以有效地提高生产效率和减少有毒废物的产生。
例如，在制备液体燃料时，低温下的加热时间缩短，可以有效减少产生有害物质的可能性。
在制备医药品时，通过调整反应温度和时间，可以确保产品的纯度和质量符合标准。
As a high school student with a keen interest in biology, Ive always been fascinated by the intricate relationship between plants and their environment. One of the most critical factors influencing plant growth and development is temperature. It plays a crucial role in various biological processes, from germination to photosynthesis, and even to the plants ability to withstand harsh conditions.My journey into understanding the impact of temperature on plants began in my schools greenhouse. Our biology teacher assigned us a project to observe the growth of different plant species under varying temperature conditions. We were to document the changes and draw conclusions about how temperature affects their growth patterns.I chose to work with a variety of seedlings, including tomatoes, sunflowers, and peas. The experiment was set up with three groups, each placed in a different temperaturecontrolled environment: one at a low temperature, one at room temperature, and one at a high temperature. Over the course of several weeks, I diligently recorded the germination rates, the speed of growth, and the overall health of the plants.What I observed was truly enlightening. The seedlings in the lowtemperature environment took longer to germinate and grew at a slower pace. Some seeds failed to sprout at all, indicating that the cold was inhibiting their metabolic processes. On the other hand, the plants in the hightemperature environment showed rapid initial growth but soon began to wilt and show signs of heat stress. Their leaves turned yellow and started to drop, suggesting that the high temperatures were causingdehydration and cellular damage.The plants in the room temperature environment, however, thrived. They germinated within a reasonable timeframe and grew steadily, showing robust health and vibrant green leaves. This group demonstrated the optimal temperature range for these species, highlighting the importance of maintaining a balance in environmental conditions.This handson experience led me to delve deeper into the subject. I learned that temperature affects the rate of photosynthesis, with most plants having a specific range of temperatures where this process is most efficient. Too cold, and the enzymes involved in photosynthesis become less active, reducing the plants ability to convert sunlight into energy. Too hot, and the plants cells can become damaged, leading to a decrease in photosynthetic efficiency and even cell death.Moreover, I discovered that temperature also influences the plants ability to adapt to its environment. Some plants, known as thermophiles, thrive in high temperatures and have developed mechanisms to prevent overheating, such as larger surface areas for heat dissipation. Conversely, psychrophilic plants are adapted to live in cold environments, often producing antifreeze proteins to prevent ice crystal formation within their cells.In agricultural settings, understanding the impact of temperature on plants is essential for optimizing crop yields. Farmers must consider factors like the planting season, the use of greenhouses, and even the selection ofcrop varieties that are best suited to the local climate. For instance, in regions with a short growing season, farmers might opt for varieties that have a shorter maturation period to ensure they can be harvested before the onset of colder weather.Furthermore, with the ongoing climate change, the study of temperatures effect on plants has become even more relevant. As global temperatures rise, plants may experience shifts in their growing seasons, altered pollination patterns, and increased vulnerability to pests and diseases. This calls for a greater understanding of plant responses to temperature fluctuations and the development of strategies to help plants adapt to these changes.In conclusion, my exploration into the world of botany has shown me that temperature is a fundamental factor in plant life. It influences everything from the most basic cellular processes to the plants ability to survive and thrive in its environment. As a high school student, I am excited to continue learning about the complexities of plant biology and hope to contribute to the ongoing research on how we can help plants adapt to the challenges posed by a changing climate.。
介绍温度句式英语作文Title: Exploring Temperature: An Insightful Composition。
Temperature is a fundamental aspect of our daily lives, influencing various phenomena and activities. From the weather outside to the comfort of our homes, understanding temperature and its implications is crucial. In this composition, we delve into the intricacies of temperature, its measurement, effects, and significance.1. Introduction to Temperature:Temperature is a measure of the degree of hotness or coldness of an object or environment. It is a vital parameter in fields such as meteorology, physics, chemistry, and everyday life. The unit of temperature commonly used in scientific contexts is the Celsius scale (°C) or Kelvin scale (K), although Fahrenheit (°F) is also prevalent in some regions.2. Measurement of Temperature:Temperature can be measured using various instruments, including thermometers, thermocouples, and infrared sensors. Thermometers, the most common instrument, typically consist of a bulb containing a temperature-sensitive liquid (suchas mercury or alcohol) connected to a calibrated scale. The liquid expands or contracts with temperature changes, indicating the temperature on the scale.3. Effects of Temperature:Temperature influences numerous natural processes and human activities. In meteorology, temperature variations drive weather patterns, affecting phenomena like precipitation, wind, and atmospheric pressure. In biology, temperature plays a critical role in determining the rangeof habitats for different species and regulating metabolic processes. Moreover, temperature impacts industrial processes, food preservation, and electronic device performance.4. Significance of Temperature Control:Maintaining optimal temperature levels is essential for various purposes. In indoor environments, proper temperature control ensures comfort and productivity. In agriculture, temperature regulation in greenhouses facilitates year-round cultivation of crops. Industriesrely on precise temperature control for manufacturing processes, ensuring product quality and efficiency. Moreover, temperature control is crucial in healthcare settings for preserving medications, vaccines, and biological samples.5. Temperature Variability and Climate Change:The Earth's climate is sensitive to changes in temperature. Human activities, such as burning fossil fuels and deforestation, have led to an increase in greenhouse gas concentrations, contributing to global warming and climate change. Rising temperatures have far-reaching consequences, including sea level rise, extreme weather events, and shifts in ecosystems. Addressing climate changerequires concerted efforts to mitigate greenhouse gas emissions and adapt to changing climatic conditions.6. Conclusion:In conclusion, temperature is a multifaceted phenomenon with significant implications for various aspects of life. From its measurement to its effects on weather, ecosystems, and human activities, temperature plays a pivotal role in shaping our world. Understanding temperature dynamics and implementing effective temperature control measures are essential for fostering sustainability and resilience in the face of ongoing environmental challenges.Through this exploration of temperature, we gain insights into the intricate workings of the natural world and the importance of responsible stewardship to ensure a harmonious coexistence with our environment.。
The impact of temperature on chemicalreactions温度对化学反应的影响化学反应是化学过程中最重要的一环，化学反应的发生往往需要一定的条件。
其中，最重要的条件之一是温度。
温度作为化学反应的重要参数之一，可以影响化学反应的速度、化学平衡等多种化学性质。
本文将从不同角度探讨温度对于化学反应的影响。
一、温度对化学反应速率的影响化学反应速率是指单位时间内反应物浓度的变化量。
在化学反应中，温度的改变对反应速率有着明显的影响。
一般而言，随着温度升高，反应速率也会随之增大。
这是因为温度升高会增加反应物分子的运动速度和撞击能量，从而使反应物分子之间发生碰撞的概率增加，化学反应的速率也就更高。
但是，当温度增加到一定程度时，反应速率将会开始减慢，这是因为高温下化学反应的物质分子速度过快，容易导致反应物分子的部分碎裂和析合，反应物降解导致反应消耗。
同时，高温下产生的自由基等反应物质越来越多，与其它反应物相互作用产生消耗。
因此，化学反应速率并非随着温度的增加而无限增加。
而是在增加至一定温度之后，开始减小。
二、温度对化学平衡的影响化学平衡是指化学反应向前和向后反应速率相等的状态。
在化学平衡下，反应物和生成物的浓度保持不变。
温度对于化学平衡的影响同样显著。
温度对酸碱反应、氧化还原反应、配位反应等各种反应类型中的平衡有着不同的影响。
以氢氧化钾的电离反应为例：KOH（aq）↔K+（aq）+OH-（aq）。
该反应的反应物中含有强碱和强酸，热力学上，其指向产物方向，即生成氢氧根离子和对应的金属离子（例如钾离子）。
在反应中添加温度，将会加快氢氧化钾的电离过程（强酸强碱易溶于热水），使九价钨酸根根离子逐渐变成威廉酸、孪晶钨酸铯等氢氧根离子、金属阳离子和水分子。
总的来说，温度对反应物的离解反应有推动效应，会促进反应物的转化为产物，使反应前进行。
在生物化学反应过程中，温度对大分子的改变，容易断裂形成小分子，此时的化学反应会增大。
The effect of temperature on reaction rateDavid A-Level 2 Background informationWhen sodium thiosulphate reacts with hydrochloric acid, a precipitate of sculpture forms. The time taken for a certain amount if sculpture to form is used to measure the reaction rate. The sodium thiosulpahte solution can be heated to different temperatures before hydrochloric acid id added, so the effect of increasing temperature can be measured.The effect of temperature increase on this reaction can be predicted using the collision theory. This theory says that for a reaction to occur, particles must collide with a certain minimum energy called the activation energy, EA. When temperature is increased, particles have increased kinetic energy and move around faster. There is therefore an increase in the frequency of collisions.Procedure:1.Measure 10 cm*cm*cm of sodium thiosulphate solution and 40 cm*cm*cm of waterinto a conical flask.2.Measure 5 cm*cm*cm of dilute hydrochloric acid in a 10 cm*cm*cm measuringcylinder3.Warm the thiosulphate solution in the flask to the requires temperature4.Put the conical flask over a piece of paper with a cross drawn on it.5.Add the acid and start timing. Swirl the flask to mix the contents .Take the initialtemperature of the mixture and record it in a table like the one below.6.Repeat the experiment using different temperatures in the range 15℃ to 65℃.Remember that one of your experiments can be carried out at room temperature andAnalysis:The diagram increase higher because the temperature increases faster.ConclusionThe reaction rate increases when the temperature increases.。
森林降温效应英文作文初中The forest cooling effect refers to the phenomenon in which forests can lower the temperature of the surrounding environment through the process of transpiration, which is the release of water vapor from the leaves of trees and plants.This cooling effect is particularly noticeable in urban areas, where the presence of trees and green spaces can help reduce the "heat island" effect caused by the concentration of buildings and pavement.In addition to lowering the temperature, forests also play a crucial role in regulating the climate by absorbing carbon dioxide from the atmosphere and releasing oxygen through the process of photosynthesis.The cooling effect of forests is not only beneficialfor the environment, but it also has a positive impact on human health and well-being. Studies have shown thatspending time in green spaces can help reduce stress and anxiety, as well as improve overall mental and physical health.Furthermore, forests provide habitat for a diverse range of plant and animal species, contributing to the overall biodiversity of an ecosystem. This, in turn, helps to maintain the balance of the natural world and ensure the stability of ecosystems.In conclusion, the forest cooling effect is an important natural process that not only helps to lower temperatures and regulate the climate, but also provides numerous benefits for both the environment and human society. It is essential to recognize the value of forests and work towards their preservation and sustainable management.。
温度英语作文Temperatures and Their ImpactTemperatures are a fundamental aspect of our daily lives, affecting everything from the clothes we wear to the activities we engage in. The range of temperatures we experience can have a significant impact on our physical and mental well-being, as well as on the natural world around us. In this essay, we will explore the various ways in which temperature can influence our lives and the world we live in.One of the most obvious ways in which temperature affects us is through the clothing we choose to wear. In colder climates, we must bundle up in warm layers to protect ourselves from the biting cold, while in warmer regions, lightweight and breathable fabrics are the order of the day. The ability to adapt our attire to the prevailing temperature is a testament to the ingenuity of the human species and our ability to thrive in a wide range of environmental conditions.Beyond the practical considerations of clothing, temperature can also have a profound impact on our physical health. Extreme heat or cold can put significant stress on the body, leading to a range of medicalconditions such as heat exhaustion, hypothermia, and even frostbite. In some cases, prolonged exposure to extreme temperatures can be life-threatening, highlighting the importance of being prepared and taking appropriate precautions.The impact of temperature is not limited to human health, however. The natural world is also highly sensitive to changes in temperature, with entire ecosystems and the species that inhabit them being affected. Shifts in temperature can lead to disruptions in the delicate balance of nature, causing species to migrate, adapt, or even become extinct. This is particularly evident in the case of climate change, where the gradual warming of the planet is leading to a host of environmental challenges that we must address.In addition to its physical impacts, temperature can also have a significant effect on our mental and emotional well-being. Many people find that their mood and energy levels are influenced by the prevailing temperature, with cold and gloomy weather often associated with feelings of sadness and lethargy, while warm and sunny days can lift our spirits and boost our motivation.This emotional connection to temperature is not just a personal quirk but is rooted in the way our brains and bodies respond to changes in the environment. For example, exposure to cold temperatures can trigger the release of hormones like cortisol, which can contribute tofeelings of stress and anxiety. Conversely, warm temperatures can stimulate the production of endorphins, which are associated with feelings of happiness and well-being.Given the multifaceted impact of temperature on our lives, it is clear that understanding and adapting to temperature variations is a crucial skill for navigating the world around us. Whether we are planning a vacation, choosing a career, or simply trying to make the most of our daily lives, being mindful of the role that temperature plays can help us make more informed and effective decisions.In conclusion, temperature is a powerful force that shapes our experiences and influences the world we live in. From the clothes we wear to the health of our planet, temperature is a constant presence that we must learn to understand and manage. By embracing the complexities of temperature and its impacts, we can better prepare ourselves for the challenges and opportunities that lie ahead.。
全球变暖效应英文作文下载温馨提示:该文档是我店铺精心编制而成，希望大家下载以后，能够帮助大家解决实际的问题。
文档下载后可定制随意修改，请根据实际需要进行相应的调整和使用，谢谢!并且，本店铺为大家提供各种各样类型的实用资料，如教育随笔、日记赏析、句子摘抄、古诗大全、经典美文、话题作文、工作总结、词语解析、文案摘录、其他资料等等，如想了解不同资料格式和写法，敬请关注!Download tips: This document is carefully compiled by theeditor. I hope that after you download them,they can help yousolve practical problems. The document can be customized andmodified after downloading,please adjust and use it according toactual needs, thank you!In addition, our shop provides you with various types ofpractical materials,such as educational essays, diaryappreciation,sentence excerpts,ancient poems,classic articles,topic composition,work summary,word parsing,copyexcerpts,other materials and so on,want to know different data formats andwriting methods,please pay attention!Global warming is a serious issue that we need to address urgently. It's not just about hotter temperaturesor melting ice caps; it's about the survival of our planet and all the living beings that call it home.Imagine a world where the sun's rays become more intense, scorching the earth's surface and making it unbearable to step outside. This is the reality we arefacing due to global warming. The greenhouse gases emitted by human activities trap heat in the atmosphere, causingthe planet to heat up at an alarming rate.The consequences of global warming are far-reaching and devastating. Rising sea levels threaten coastal communities, leading to the displacement of millions of people. Extreme weather events such as hurricanes and droughts become more frequent and intense, causing widespread destruction andloss of life. The delicate balance of ecosystems is disrupted, leading to the extinction of countless species.It's not just the environment that is affected by global warming; it's also our health. Heatwaves become more frequent, putting vulnerable populations at risk of heatstroke and other heat-related illnesses. Air pollution worsens as higher temperatures increase the formation of smog and the spread of allergens. The quality of our food and water is compromised, leading to food shortages and water scarcity.Addressing global warming requires a collective effort from individuals, governments, and businesses. We need to reduce our carbon footprint by adopting sustainable practices such as using renewable energy sources and conserving water. Governments should implement stricter regulations on emissions and invest in green technologies. Businesses should prioritize sustainability and invest in research and development to find innovative solutions to combat global warming.Education also plays a crucial role in tackling global warming. We need to raise awareness about the causes andconsequences of global warming and empower individuals to take action. By educating the younger generation about the importance of protecting the environment, we can create a sustainable future for all.In conclusion, global warming is a pressing issue that requires immediate action. The consequences of inaction are dire, with catastrophic effects on our planet and its inhabitants. We must all come together and make changes in our daily lives, support policies that promote sustainability, and educate others about the urgency of addressing global warming. Our future depends on it.。
怎样介绍温度英语作文Title: Understanding Temperature: A Comprehensive Introduction。
Temperature is a fundamental aspect of our daily lives, influencing everything from the weather to the behavior of substances. In this essay, we will delve into the conceptof temperature, exploring its definition, measurement, effects, and significance in various contexts.Definition and Measurement:Temperature is defined as the degree of hotness or coldness of a body or environment. It is a measure of the average kinetic energy of the particles in a substance. The most common unit of temper ature is degrees Celsius (°C), although other units such as Fahrenheit (°F) and Kelvin (K) are also used.Temperature can be measured using various instruments,such as thermometers. Traditional liquid-in-glass thermometers, digital thermometers, and infrared thermometers are among the tools commonly employed to determine temperature accurately.Effects of Temperature:Temperature exerts significant influence on both living organisms and non-living systems. In living organisms, temperature plays a crucial role in biological processes. For instance, human body temperature must remain within a narrow range (approximately 36.5°C to 37.5°C) for optimal physiological functioning. Extreme deviations from this range can lead to health problems, such as hypothermia or hyperthermia.In non-living systems, temperature affects the physical properties of substances. For example, changes in temperature can cause solids to melt into liquids and liquids to vaporize into gases. This phenomenon is evident in everyday experiences, such as the melting of ice cream on a hot day or the boiling of water when heated.Significance in Various Contexts:1. Weather and Climate: Temperature plays a centralrole in determining weather conditions and climate patterns. It influences phenomena such as precipitation, wind patterns, and the formation of weather systems like hurricanes and tornadoes. Climatologists use temperature data to analyze long-term climate trends and predict future climate changes.2. Industry and Engineering: In industrial processes, temperature control is essential for ensuring the quality and efficiency of manufacturing operations. From food processing to semiconductor fabrication, precisetemperature regulation is vital for achieving desired outcomes and maintaining product integrity.3. Environmental Impact: Temperature variations have significant implications for the environment. Global warming, driven by increased atmospheric concentrations of greenhouse gases, has led to rising temperatures worldwide.This phenomenon has far-reaching consequences, includingthe melting of polar ice caps, sea-level rise, and changesin ecosystems and biodiversity.4. Medicine and Healthcare: Temperature monitoring is crucial in medical settings for diagnosing and treating various health conditions. Healthcare professionals use thermometers to measure body temperature and identify fever, a common symptom of infections and illnesses. Additionally, temperature-controlled environments are necessary forstoring medications, vaccines, and biological samples.Conclusion:In conclusion, temperature is a fundamental conceptwith widespread implications across diverse fields and disciplines. Whether influencing the weather, driving industrial processes, shaping environmental dynamics, or impacting human health, temperature exerts a profound influence on our lives. By understanding the principles of temperature and its effects, we can better navigate and adapt to the dynamic world around us.。
Effect of temperature on AOB activity of a partial nitritation SBR treating landfill leachate with extremely high nitrogen concentrationJ.Gabarróa ,⇑,R.Ganiguéa ,b ,F.Gich c ,M.Ruscalleda a ,M.D.Balaguer a ,J.Colprim aaLEQUIA,Institute of the Environment,University of Girona,Campus Montilivi,E-17071Girona,Catalonia,SpainbAdvanced Water Management Centre,Building 60,Research Road,The University of Queensland,St.Lucia,Brisbane,QLD 4072,Australia cGroup of Molecular Microbial Ecology,Institute of Aquatic Ecology (UdG),Campus Montilivi,E-17071Girona,Catalonia,Spainh i g h l i g h t s"Successful partial nitritation at 25and 35°C treating leachate at 6g N-NH 4+L À1."A kinetic model was applied to study the influence of FA and FNA inhibition on AOB."FNA was the main source of AOB inhibition at 25°C."At 35°C,AOB inhibition was due to the combined action of FA and FNA."Two AOB phylotypes were found during stable state at 25and 35°C.a r t i c l e i n f o Article history:Received 16July 2012Received in revised form 4September 2012Accepted 5September 2012Available online 13September 2012Keywords:Partial nitritation Free ammonia (FA)Free nitrous acid (FNA)Landfill leachate Temperaturea b s t r a c tThis study investigates the effects of temperature on ammonia oxidizing bacteria activity in a partial nit-ritation (PN)sequencing batch reactor.Stable PN was achieved in a 250L SBR with a minimum operating volume of 111L treating mature landfill leachate containing an ammonium concentration of around 6000mg N-NH 4+L À1at both 25and 35°C.A suitable influent to feed an anammox reactor was achieved in both cases.A kinetic model was applied to study the influence of free ammonia (FA),the free nitrous acid (FNA)inhibition,and the inorganic carbon (IC)limitation.NH 4+and NO 2Àconcentrations were sim-ilar at 25and 35°C experiments (about 2500mg N-NH 4+L À1and 3500mg N-NO 2ÀL À1),FA and FNA con-centrations differed due to the strong temperature dependence.FNA was the main source of inhibition at 25°C,while at 35°C combined FA and FNA inhibition occurred.DGGE results demonstrated that PN-SBR sludge was enriched on the same AOB phylotypes in both experiments.Ó2012Elsevier Ltd.All rights reserved.1.IntroductionUrban landfill leachate characteristics vary greatly depending on factors such as seasonal weather variation,waste type and com-position,and landfill age,among others.The biological treatment of landfill leachate mainly focuses on organic matter and ammo-nium removal,and is mostly carried out through conventional autotrophic nitrification–heterotrophic denitrification.However,when treating mature landfill leachates characterized by low C/N ratio,high aeration and an external carbon source are required,entailing high operational costs.The anaerobic ammonium oxidation (anammox)process is a more sustainable alternative for the treatment of wastewaters con-taining high nitrogen loads and low biodegradable organic matter (Van Dongen et al.,2001).As a result,the feasibility of this process for landfill leachate treatment has previously been studied at low and high ammonium concentration (Ganiguéet al.,2009;Ruscalleda et al.,2008;Siegrist et al.,1998).The anammox process requires a previous partial nitritation (PN)in order to produce a NO 2À:NH 4+molar ratio of 1.32(Strous et al.,1999).PN systems aim to out-compete nitrite oxidizing bacteria (NOB)to achieve nitrite accumulation by ammonia oxidizing bacteria (AOB)and with the startup as a key period to inhibit and/or washout NOB.Several controlling factors can be applied to out-compete NOB and ensure nitrite accumulation,such as short sludge retention time (SRT),pH,dissolved oxygen (DO),free ammonia (FA)or free nitrous acid (FNA)inhibition or temperature (Ahn et al.,2011,Liang and Lu,2007,Van Dongen et al.,2001).Focusing on temper-ature,PN reactors are usually operated between 30and 35°C,be-cause at these values AOB have higher growth rates than NOB (Hellinga et al.,1998).However,temperature also affects the chemical equilibria of ammonium-free ammonia (FA)and nitrite-free nitrous acid (FNA),which can play a critical role in partial nit-ritation.High FA and FNA concentrations affected by temperature0960-8524/$-see front matter Ó2012Elsevier Ltd.All rights reserved./10.1016/j.biortech.2012.09.011Corresponding author.Tel.:+34972183244;fax:+34972418150.E-mail addresses:jgabarro@lequia.udg.cat ,jgabarro@ (J.Gabarró).may reduce or even inhibit not only NOB but also AOB activity, whereas a reduction of the inhibition pressure could allow NOB development(Anthonisen et al.,1976;Vadivelu et al.,2007a;Van Hulle et al.,2007).Landfill leachates are usually stored for long periods in ponds at ambient temperature prior to their treatment.Hence,reactors may require heating to ensure suitable operating conditions, increasing the treatment costs.Several studies demonstrated the suitability of the anammox process at low temperatures (20–25°C,(Vazquez-Padin et al.,2011;Yang et al.,2011)).With regard to PN reactors,they have been operated in different configurations and temperatures(Jeanningros et al.,2010;Peng et al.,2008;Vazquez-Padin et al.,2011).However,few studies have investigated the impact of temperature on AOB activity taking into account FA/FNA inhibition as well as IC limitation.Egli et al. (2003)assessed the impact of slightly different temperature(25 and30°C)and pH(7and7.5)on the startup of nitritation reactors with different configurations and fed with synthetic media (700–1000mgN LÀ1)and sludge digester supernatant,reporting successful and stable nitrite build-up for all conditions,with different AOB phylotypes and temperature playing a minor impact. According to Vejmelkova et al.(2012),ammonium concentration has a more significant role than temperature on the dominant AOB phylotype.However,when treating landfill leachate containing an extremely high concentration of nitrogen(up to6g N-NH4+LÀ1), temperature could have a more critical role because of its impact on FA and FNA equilibria.In this light,the main goal of this study was to establish the temperature effects on a PN reactor taking into account FA/FNA inhibition and inorganic carbon limitation at25and35°C treating landfill leachate with an extremely high concentration of nitrogen concentration,as a prior step for an anammox reactor.The study also aimed to assess the change in the AOB community during the startup and stabilization phase.2.Methods2.1.Raw leachateRaw leachate was collected from the Corsa urban landfill site (Reus,Catalonia,Spain).It was conditioned and stored in a300L tank.Leachate conditioning was obtained by adding sodium bicar-bonate to ensure an influent bicarbonate:ammonium molar ratio of1.14(Ganiguéet al.,2009).The landfill leachate characterization after bicarbonate adjustment is presented in Table1.2.2.Experimental set-up and operationTwo successive experiments were conducted in a250L pilot scale sequencing batch reactor(SBR).One experiment was per-formed at25°C and the other at35°C.The reactor had a minimum operating volume of111L and was water-jacketed,allowing the mixed liquor temperatures to be controlled by a thermostated water bath.The reactor was equipped with a mechanical stirrer to ensure complete mixing.Aeration was carried out by air diffus-ers(Magnum,from OTT System GmbH&CO.,Germany)placed at the bottom part of the reactor.The pilot plant operation was super-vised by a control system with own-developed software.The pH, oxidation reduction potential(ORP),DO and temperature were monitored using CPF81,CPF82and OXYMAX-W COS-41probes (Endress+Hauser,Germany).Control actions were transmitted by an interface card(PCI-885from Advantech,USA)connected to a re-lay output board,which controlled the on/off switch of electrical devices.The partial nitritation SBR(PN-SBR)cycle had a total duration of 1440min.The cycle was divided into14sub-cycles of100min each,a20-min settling phase and a20-min draw phase.Every sub-cycle consisted of10min of aerobic feeding and90min of aer-obic reaction.No purge phase was applied because of the high sus-pended solids concentration in the effluent(400–700mgVSS LÀ1).Seed sludge for the experiments was obtained from the Sils-Vidreres urban wastewater treatment plant(WWTP;Sils, Catalonia,Spain).Reactors were started at an specific nitrogen loading rate(sNLR)of0.2kgN kgVSSÀ1dÀ1,which was progres-sively raised during the study by increasing the daily influentflow. At stable state,hydraulic retention time(HRT)was about4.5and 12.0days at25and35°C,respectively.Additionally,during stable phase,the reactor suspended solids concentration was1306±620mgVSS LÀ1.The temperature was maintained at25and35°C, depending on the experiment.The DO set-point wasfixed at a minimum value of2mgO2LÀ1and pH was controlled below a maximum of8.0by the addition of hydrochloride acid(1M).2.3.Analytical methodsThe concentrations of ammonium(NH4+),inorganic carbon(IC), nitrite(NO2À),nitrate(NO3À),total Kjeldahl nitrogen(TKN), chemical oxygen demand(COD),biochemical oxygen demand (BOD x),and total and volatile suspended solids were determined according to standard methods(APHA,2005).Conductivity and pH were measured with a conductometer(EC-Meter Basic30+ from Crison,Spain)and a pH meter(pH-Meter Basic20+from Crison,Spain)respectively.2.4.Molecular techniques2.4.1.DNA isolationDNA was extracted from1ml of mixed liquor samples collected from the reactors at different time points with the FastDNAÒSpin kit for soil(MP Biomedicals,Solon,OH,USA),in accordance with the manufacturer’s instructions.DNA concentration and purity were determined in a Nanodrop ND-1000UV–Vis spectrophotom-eter(Nanodrop,Wilmington,DE,USA).2.4.2.Real-time PCR quantification of total AOBGene copy numbers for the ammonia-oxidizing bacteria(AOB) 16S rRNA gene were determined by quantitative real-time PCR (qPCR)amplification on DNA extracts from both reactors at differ-ent time points.All qPCR assays were performed in a7500real-time PCR system(Applied Biosystems,Carlsbad,CA,USA)on MicroAmp optical96-well reaction plates covered with optical caps(Applied Biosystems,Carlsbad,CA,USA)using the primers CTO189f/RT1r(Table2).Primers CTO189A/Bf and CTO189Cf were used at a2:1ratio.Inhibition of16S rRNA gene amplification was checked for each DNA extract using the environmental clone R450RT9(accession number FR875176)cloned into the TOPO TA vector(Invitrogen,Carlsbad,CA,USA)as internal control.TheTable1Raw leachate characteristics(n=38samples).Compound Units Mean±dAmmonium(NH4+)mgN LÀ15975±213 Nitrite(NO2À)mgN LÀ10.00±0.00 Nitrate(NO3À)mgN LÀ1<2Total Nitrogen(TN)mgN LÀ16317±251 Inorganic carbon a mgC-HCO3ÀLÀ15488.9±512.1 Chemical oxygen demand(COD)mgO2LÀ13921±1264 Biochemical oxygen demand(BOD5)mgO2LÀ1123±88.9 Conductivity(EC)l S cmÀ174060±2816 pH–8.54±0.12a Adjusted by Na HCO3addition to1.14mol C-IC/mol N-NH4+.284J.Gabarróet al./Bioresource Technology126(2012)283–289clone sequence R450RT9was amplified with the primers T3and T7 binding toflanking regions in the TOPO TA vector according to the manufacturer’s instructions(data not shown).All the reaction mixtures(20l l)contained10l l of SsoFast EvaGreen supermix (Bio-Rad,Hercules,CA,USA),2l l of template DNA(10ng),0.4l l each of primer CTO189f and RT1r at afinal concentration of 300nM(Biomers,Ulm,Germany),2l l of bovine serum albumin (10mg/ml)(Sigma–Aldrich,Steinheim,Germany),and 5.2l l of molecular biology-grade water(Sigma–Aldrich,Steinheim, Germany).The qPCRs were run as follows:enzyme activation for 30s at95°C,followed by35cycles of5s at95°C(denaturaliza-tion)and30s at60°C(annealing and extension).Thefluorescence signal was read in each cycle after the elongation step at80°C for 32s for accurate stringent product quantification.All reactions were run in duplicates with standard curves spanning from101 to108copies of DNA of the16S rRNA gene.Standard curves were obtained after serial dilutions of previously titrated suspensions of the16S rRNA gene amplified by conventional PCR from the envi-ronmental clone R450RT9and further purification(6Minute Mini Plasmid Prep Kit™MoBio,Carlsbad,CA)and quantification using Nanodrop.Prior to qPCR the environmental clone R450RT9was linearized by enzymatic digestion with SpeI to avoid super coiling during real-time PCR which could potentially influence the PCR yield.The PCR efficiency(E)of the standard curve calculated according to E=10(À1/slope)À1was90.93%with R2values of >0.99and an equation slope ofÀ3.32(Kubista et al.,2006).The specificity of reactions was confirmed by melting-curve analyses and by agarose gel electrophoresis to identify unspecific PCR prod-ucts,such as primer dimers or gene fragments of unexpected length(data not shown).2.4.3.PCR amplification and DGGEfingerprinting16S rRNA gene sequences for the ammonia-oxidizing bacteria (AOB)were amplified from four mixed liquor samples at different reactor operational conditions;two at25°C(days35and42)and two at35°C(days73and79).PCR amplifications were performed in a GeneAmp PCR System9700(Applied Biosystems,Foster City, CA,USA)using the forward primers CTO189A/Bf and CTO189Cf (2:1)and the reverse primer CTO654r as previously described (Limpiyakorn et al.,2005).Forward primers contained a40-bp-long GC clamp at their50end in order to obtain stable melting behavior of the generated DNA fragments during the subsequent DGGE.A constant amount of35ng of genomic DNA from each ex-tract was used for all PCR reactions to avoid differences in the amplification yield.PCR products were generated in duplicates in a reaction volume of50l l,mixed,concentrated to35l l and separated by DGGE in6%(wt/vol)polyacrylamide gels containing a linear gradient of30–80%denaturant using an Ingeny phorU system(Ingeny International BV,Goes,The Netherlands)as de-scribed earlier(Llirós et al.,2010).All identified discrete and clear bands were excised from the gels and rehydrated in20l l of 10mM Tris–HCl buffer(pH7.4).DNA was eluted after incubation at65°C for2h and amplified using the corresponding primer pairs(without a GC clamp)cited above using the same PCR con-ditions.PCR products were cleaned and sequenced as described by Gich et al.(2005).Sequences of the closest relatives were retrieved from the GenBank database by employing the BLAST alignment tool at the National Center for Biotechnology Informa-tion(NCBI).2.5.Nucleotide sequence accession numbers16S rRNA gene sequences obtained in this study are available at the EMBL database under accession numbers JQ673432through JQ673439.3.Calculations3.1.Free ammonia and free nitrous acidThe values of the free ammonia(FA)and the free nitrous acid (FNA)concentrations were calculated as a function of pH,temper-ature and total ammonium as nitrogen(TAN),for FA,or total nitrite (TNO2),for FNA(Eqs.(1)and(2);Anthonisen et al.,1976).FAðmgNÁLÀ1Þ¼TAN1þ10ÀpHK e;NH3where;K e;NH3¼À6344e273þTð1ÞFNAðmgNÁLÀ1Þ¼TNO21þK e;HNO210ÀpHwhere;K e;HNO2¼À2300ð2ÞAccording to Eqs.(1)and(2),it is shown that temperature has an inverse effect on the FA and FNA equilibrium constants.3.2.Observed and maximum nitrite production rateThe observed nitrite production rate(NPR obs)was calculated as a function of daily influentflow(Q d;(L dÀ1)),nitrite[NO2À] (mgN LÀ1),nitrate[NO3À](mgN LÀ1)and maximum volume of the reactor(V max;(L))according to Eq.(3).NPR obs¼QdÁ½NOÀ2þQdÁ½NOÀ3V maxðmgNÁLÀ1ÁdÀ1Þð3ÞThe specific nitrite production rate(sNPR)was calculated as a function of NPR obs and the total AOB concentration[AOB tot] (cells LÀ1)as given in Eq.(4).sNPR obs¼NPR obsÁ1010½AOB totð1010mgNÁcellsÀ1ÁdÀ1Þð4ÞOn the other hand,a maximum specific nitrite production rate (sNPR max)(mgN1010cellsÀ1dÀ1)was calculated from Eq.(4)fol-lowing the kinetic expression and constants previously described in literature(Ganiguéet al.,2007),which describes the sNPR obs to be dependent on the NPR max,FA and FNA inhibition(modeled as Monod terms)as well as bicarbonate concentration(modeled as a sigmoidal term).sNPR max¼sNPR obsK AOBI;NH3K AOBI;NH3þNH3ÁKAOBI;HNO2K AOBI;HNO2þHNO2Áe HCOÀ3=ae HCO3ðÞ=aðÞþ1ð5ÞTable2Group-specific PCR primers used to amplify the16S rDNA gene sequences from ammonia oxidizing bacteria.Primer Specificity Sequence(50–30)ReferencesCTO189A/Bf b-Proteobacteria(AOB)GGAGRAAAGCAGGGGATCG Kowalchuk et al.(1997) CTO189Cf b-Proteobacteria(AOB)GGAGGAAAGTAGGGGATCG Kowalchuk et al.(1997) RT1r AOB CGTCCTCTCAGACCARCTACTG Hermansson and Lindgren(2001)J.Gabarróet al./Bioresource Technology126(2012)283–2892854.Results and discussion 4.1.Startup and operationThe PN-SBR was inoculated twice,in a different time period,with 200L of mixed liquor (1000mgVSS L À1)from a conventional urban WWTP that removed organic matter and nitrogen.The PN-SBR was operated at two different temperatures,25and 35°C.The reactor was solely fed with raw leachate (Table 1)from the beginning of the startup.Partial nitritation (PN)was achieved by bicarbonate limitation on AOB.Ganiguéet al.(2009)specially re-ferred to the stoichiometric influent molar ratio HCO 3À:NH 4+of 1.14to achieve the desired effluent molar ratio NO 2À:NH 4+of 1.32to feed an anammox reactor.Fig.1presents the evolution of the main nitrogen compounds and the specific nitrogen loading rate (sNLR)over time for both experiments (25and 35°C).For both experiments,the initial sNLR was set at 0.2kgN ÁkgVSS À1d À1,and was progressively increased to values up to 0.8–1kgN ÁkgVSS À1d À1at the operational phase.A slower sNLR increase was applied at 25°C than at 35°C.During the entire study,the influent concentration of ammonium was about 6000mg N-NH 4+L À1.Nitrate was produced during the first 10days,reaching a maximum concentration of around 250mg N-NO 3ÀL À1for both the 25°C and the 35°C experiments.After the 10th day,NO 3Àeffluent concentration steadily decreased,reaching values below 50mg N-NO 3ÀL À1at the end of the experi-ments.With regards to NO 2À,it accumulated in the system from day 0in both experiments,reaching stable concentration values –around 3500mg N-NO 2ÀL À1–from day 22nd and 18th,at 25and 35°C,respectively.The ammonium concentration in the efflu-ent was linked to nitrite/nitrate production.In this sense,the NH 4+effluent concentration increased from day 0,showing a trend sim-ilar to the nitrite concentration in both conditions.After the start-up period,the reactor was operated in a stable manner at both temperatures.Table 3summarizes the most important features of the reactor during the operational period.Effluent characteristics were similar for both experiments (NH 4+2700mgN L À1,NO 2À3500mgN L À1,NO 3À35mgN L À1),and were mainly governed by the HCO 3À:NH 4+influent molar ratio.It is also remarkable that the nitrate concentration was lower than 1%of the total influent nitrogen for both conditions,demonstrating a good performance of the PN process.High nitrite accumulation and very low nitrate formation were due to AOB growth and NOB out-competition.As previously stated,NOB wash-out or inhibition can be achieved by several factors.Gi-ven the extremely high ammonium content of the landfill leachate,FA and FNA concentrations were the main factors for NOB activity suppression in this study according to previous literature (Anthon-isen et al.,1976;Vadivelu et al.,2007b ).Additionally,AOB activity may also be negatively affected by both FA and FNA inhibition.This was further explored in Fig.2,which shows the evolution of max-imum FA and FNA concentrations as well as daily average pH at 25and 35°C.FA and FNA concentrations were calculated according to Eqs.(1)and (2).As can be seen in Fig.2,in both experiments,the average pH was around 7,being slightly higher at 35°C (7.00vs.7.33,at 25°C and 35°C respectively).During the startup period,FA and FNA concentrations increased rapidly together with ammonium and nitrite concentrations (Fig.1).At day 10th,the FA concentra-tion was 10and 25mg N-NH 3L À1at 25and 35°C,whereas the FNA concentration was about 0.2mg N-HNO 2L À1for both conditions.When looking at the operational performance levels,the FA and FNA concentrations were significantly different.At 25°C,FA and FNA were on average 20.76±4.23mg N-NH 3L À1and 0.47±0.09mg N-HNO 2L À1,respectively.On the other hand,at 35°C FA was much higher,122.92±27.23mg N-NH 3L À1,whereas FNA levels were 0.12±0.02mg N-HNO 2L À1,lower than at 25°C.This opposite behavior of FA and FNA with regard to temperature is observed by analyzing Eqs.(1)and (2):FA would be higher as temperature increased,whereas FNA would be lower.In this sense,Table 3Main parameters of the PN-SBR operation during the stable period.ParameterUnitsTemperature 25°C35°CsNLR(KgN kgVSS À1d À1)0.81±0.110.84±0.24HCO 3À:NH 4+inf.(mol HCO 3Àmol À1NH 4+) 1.16±0.06 1.12±0.06NH 4+eff (mgN L À1)2725.9±153.22629.9±123.4NO 2Àeff (mgN L À1)3719.2±174.53245.5±115.7NO 3Àeff(mgN L À1)41.9±25.025.8±3.0NO 2À:NH 4+eff.(molNO 2Àmol À1NH 4+) 1.33±0.08 1.23±0.07IC eff(mgC-HCO 3ÀL À1)52.4±22.945.7±10.6286J.Gabarróet al./Bioresource Technology 126(2012)283–289it was expected tofind lower FA values at25than at35°C,and the opposite would be expected regarding FNA levels.Having the same amount of nitrite and ammonium,different levels of FA and FNA would be found in function of the operational temperature.4.2.AOB community during stable operationStringent conditions such as FA and FNA concentrations lead to a decrease in diversity of the AOB community(Gregory et al., 2010).Therefore,the AOB members sludge composition present in the reactors was analyzed during the operational period.The seeding sludge was collected and analyzed by PCR as previously re-ported(Ganiguéet al.,2009).Fingerprint patterns were generated by DGGE from two replicas of DNA samples from both25°C(days 35th and42nd)and35°C(days73rd and79th).Identicalfinger-prints were identified for all four samples containing only two melting types(Fig.3).For the samples at35°C,the second melting type showed lower signal intensity than for the25°C samples.It must be mentioned that the amount of DNA used in the PCR reac-tion was the same(35ng)and PCR reactions were performed in duplicates in the same PCR run.Xie et al.(2012)found that media and temperature are the two main determinants of bacterial com-munity structure.Thus,the most plausible explanation for our findings would be that the presence of the second melting type is more abundant at25°C because of the lower temperature and, consequently,lower FA and higher FNA concentrations.Another plausible explanation would be the wash-out of the second melt-ing type at35°C,however,this seems unlikely because HRT was higher at35°C.In this sense,Tan et al.(2008)reported similar observations,and two AOB phylotypes were detected in the sludge of a reactor with nitrite concentration up to500mM.Nevertheless, a longer operational period would be needed to have a clear under-standing of the long-term evolution of the community and eluci-date the main reason for the differential behavior at25and35°C.All eight discrete DGGE bands were excised,the DNA was elutedand sequenced.16S rRNA partial gene sequences(382bp)corre-sponding to bands1,2,3and4were100%identical whereas those corresponding to bands5,6,7and8showed a similarity ranging from98.1to99.4%,indicating a unique AOB phylotype(A or B, Fig.3)for each melting type.In addition,both melting types showed a similarity of95.0to95.5%.Since a sequence similarity >97%corresponds to the same bacterial species(Stackebrandt and Goebel,1994),these16S rRNA gene sequences may represent at most two bacterial species present in all the analyzed samples. On the contrary,both phylotypes could actually originate from multiple ribosomal RNA(rrn)operons that are present in the same bacterium.In the present case,however,this appears unlikely because Nitrosomonas europaea ATCC19718and Nitrosomonas eutropha C91both contain one single rrn operon copy number according to the rrn database(Lee et al.,2009)and,therefore,each phylotype would correspond to different bacteria.It is generally assumed that AOB possess one copy of the16S rRNA per genome based on the16S rRNA gene copies found in AOB(Aakra et al., 1999).The phylogenetic analysis from the present study of thefinger-prints revealed that the phylotype A was100%identical to an uncultured Nitrosomonas sp.(FM997776)repeatedly identified in a partial nitrification reactor treating mature leachates at35°C in long-term monitoring during the years2008and2009(Ganigué12345678SybrGold-stained gel of ammonia-oxidizingbp)separated by DGGE.Numbersthat were excised and sequenced.same position harbored highly similarrespectively).The percentages on thedenaturant.ST,DGGE band positionJ.Gabarróet al./Bioresource Technology126(2012)283–289287et al.,2009).Since the inoculum of that study contained5different AOB phylotypes,our results clearly demonstrates a selection of two phylotypes.With regards to phylotype B,it showed99.0%sim-ilarity with an uncultured bacterium(HQ228600)identified in a nitrification reactor for treating high-strength nitrogen wastewater (Ahn et al.,2011).The closest cultivated representative to phylo-type A and B was N.europea strain ATCC25978with a similarity of95%and99%,respectively.Ammonium influent concentration is a more important parameter than temperature for AOB phylo-type selection(Egli et al.,2003;Gregory et al.,2010;Vejmelkova et al.,2012).In this sense,our results are in total accordance with previous related literature.4.3.PN-SBR sludge activityDGGE results demonstrated that PN-SBR sludge was enriched on the same AOB phylotypes in both25and35°C experiments. In this sense,copy numbers of16S rRNA gene for AOB were deter-mined by qPCR for both performances(25°C:days2nd,14th,22nd, 29th,36th;and35°C:days8th,17th,25th,38th,44th,57th,63rd, 73rd).The cell numbers of total AOB were calculated assuming one operon copy of16S rRNA gene per genome(Aakra et al.,1999).To-tal numbers of AOB were used to calculate the sNPR obs from Eq.(4). Results are depicted in Fig.4.During the startup,sNPR obs progressively increased,showing a similar trend in both experiments.The experiment at25°C ended after45days,it is unclear whether sNPR obs would have been fur-ther increased in time when the reactor would have been operated longer.The highest sNPR obs obtained during the stable operation phase were9.1Â10À10and14.3Â10À10mgNÁcellsÀ1dÀ1at25and35°C,respectively.High FA and FNA concentrations in the mixed liquor(Fig.2) may have had a negative impact on AOB activity.Therefore,the theoretical maximum sNPR(sNPR max)was calculated according to Eq.(5).The sNPR max calculated was about32.3Â10À10-mgNÁcellsÀ1dÀ1at25°C,while it was46.7Â10À10mgNÁcellsÀ1dÀ1 at35°C.The ratio between sNPR obs and sNPR max for both experi-ments is shown in Fig.5.During the startup phase(up to day20th),both experiments de-picted a decreasing trend of the ratio sNPR obs:sNPR max(Fig.5)from an initial value of0.55due to the increase in FA and FNA concen-trations.However,the ratio was kept constant during the stable operational period,remaining at0.28and0.31at25and35°C respectively.These low ratios at both temperatures were caused by FA and FNA inhibition.Also,IC limitation influenced these low ratios.In order to gain further insight on the impact of IC limitation and FA/FNA inhibition on AOB activity at two different temperatures(25and35°C),the contribution on the AOB activity reduction of each kinetic term was calculated based on Eq.(5) during the stable operational period.Results are presented in Table4.IC limitation played a similar role at both temperatures, with AOB activity reduction at around45%.There was,however, a differential behavior in the impact of FA and FNA on AOB. Whereas FNA inhibition term had a contribution on the AOB activity reduction of49.9%and21.6%AOB at25and35°C,respec-tively,FA only had a significant effect at35°C(22.1%).Analyzing both experiments(25and35°C),independently of the reactor temperature,the sNPR obs were much lower than the theoretical sNPR max(about30%,Fig.5).These lower sNPR were mainly due to two different factors:IC limitation and FA/FNA inhi-bition.PN was achieved by IC limitation to control the conversion of the influent nitrogen oxidation to nitrite to50%.This also af-fected AOB activity due to IC limitation(Guisasola et al.,2007; Wett and Rauch,2003).Taking into account these limiting condi-tions,in the present study,IC limited nitrite production by42% and49%at25and35°C respectively(Table4).These results were expected due to the limitation of the nitritation by bicarbonate concentration.The other factor that influenced AOB activity was the FA/FNA inhibition.Although ammonium and nitrite effluent concentra-tions were similar at25and35°C performances,the FA and FNA concentrations differed due to their temperature dependence. Therefore,FA and FNA temporal evolutions were different for each experiment as presented in Fig.2.AOB activity is inhibited by both FA and FNA(Anthonisen et al.,1976;Ganiguéet al.,2009;Vadivelu et al.,2007a).In this sense,FNA was the main inhibiting factor affecting AOB activity at25°C together with IC limitation.FNA sta-bilized around0.5mg N-HNO2LÀ1and the calculated contribution of FNA inhibition was about50%(Table4).Our results were similar to previous work where the same FNA concentration contributed to a50%of activity depletion of Nitrosomonas pure culture (Vadivelu et al.,2007b).On the other hand,the main cause ofTable4Contribution of FA,FNA inhibition and IC limitation kinetic terms to AOB activitydepletion at25°C and35°C.FA inhibition(%)FNA inhibition(%)IC limitation(%)1ÀKAOBI;NH3KI;NH3þNH31ÀKAOBI;HNO2KI;HNO2þHNO21Àe HCOÀ3ðÞ=aðÞeÀ3þ1Temperature Mean(%)±d(%)Mean(%)±d(%)Mean(%)±d(%)25°C 4.9±0.949.9±0.642.2± 1.535°C22.1± 2.321.6± 1.749.4±0.7 288J.Gabarróet al./Bioresource Technology126(2012)283–289。
