Chemical and Solar Adaptations

to Winter

The density of pure water ice at 0°C is 0.9168 g/ml, nearly 9% lighter than liquid water at

0°C, which has a density of 0.99987 g/ml. It is enough to keep ice floating on top of water

and allows aquatic organisms to survive the winter. This drop in density occurs because the hydrogen bonds in water create an open hexagonal lattice, leaving space between the

molecules.

Ice Crystal Snow

Colligative Properties

• Some of the properties of solutions do not Some of the properties of solutions do not depend on the amount and type of solute depend on the amount and type of solute present in solution.present in solution.– Ie food coloring doesn’t affect the boiling point Ie food coloring doesn’t affect the boiling point

of water (much)of water (much)

• Properties that depend on the Properties that depend on the concentration of solute particles but not on concentration of solute particles but not on their identity are called Colligative their identity are called Colligative properties.properties.

Electrolyte and non Electrolyte

• Electrolytes are substances that dissolve in water to give a solution that conducts an electric current– Sports drinks and salt water– Ionic compounds are usually strong

electrolytes because they separate completely in water

– Covalent compounds can be strong, weak or non electroyltes

• Non-electrolytesNon-electrolytes: a liquid or solid : a liquid or solid substance that does not allow the flow substance that does not allow the flow of an electric current, either in solution of an electric current, either in solution or in its pure state, such as water or or in its pure state, such as water or sucrose.sucrose.

• Nonvolatile substance is one that has Nonvolatile substance is one that has little tendency to become a gas under little tendency to become a gas under existing conditionsexisting conditions

What about Electrolytes?

• Electrolytes break apart into ions. Each ion has an effect on boiling point and freezing points. If a solution has more or less ions it will change the boiling points and melting points even more.

Plants and Animals in Winter

• Plants and Animals must adapt to the coming of winter by:– Behavioral Adaptations

• Bird Migration

• Bear Hibernation

– Physical Adaptations• Deciduous trees drop leaves in the fall

• Wood frogs and Cope’s grey tree frogs survive by freezing solid

Physical Adaptations

• With most physical adaptations to winter there is an important chemical adaptation– This is in response to:

1) Water freezing which, causes

2) Ice crystals to form damaging cells

• Both plants and animals have a variety of chemical adaptations

Productivity

• Energy pyramid is skewed in winter– Primary producers shut down– Movement and travel require more energy– Exposure cause energy drain.– Food production low– Need for more food is high

Plant sources of heat and loss

The complex system of acclimation carbonic anhydrase II

Catalizes hydrationOf Carbon Dioxide

Kinases enzyme catalyzes phosphorous

cold response and improves freezing tolerance of the transgenic plants regulator

CBF – blood flow or fluid

Protein binding gene

Radical Death gene Melatonin induced

Histomine usually in plant seed coating

Deciduous Trees, Abscisic Acid, Fructose, Glucose and

Sucrose

Sugar Maple

• Taking a sugar substance from a tree can be used to create maple syrup

• The maple syrup aqueous solution boils at 219 degree F

• That is 7 degree higher than water.

• It also depresses the freezing point

Deciduous Trees• Deciduous trees start producing abscisic

acid– Due to reduction in photoperiod (seasonal)– In response to reduction of water– Because of drop in nutrients for the tree

Abscisic Acid

• Is released in response stress• The main cause of abscission (leaf loss) in

deciduous trees• Inhibits cell division in the cambian (why tree

rings in winter are narrower)• Hardens cell membranes to help protect against

ice crystals

Growth inhibiting substance as Abscisic acid or Abscissin II, which was once called as ‘Dormin’. Besides ABA, plants also contain other natural growth inhibitors such as Coumerin, Ferulic acid, Para ascorbic acid, phaseic acid, violoxanthin, etc. In addition, plant chemists have identified some synthetic growth inhibitors. Ex. 2,3,5 tri iodo-benzoic acid, morphactins, caproic acid, phenyl propionic acid, Malic hydrazide, etc.

ABA RESPONSE CHART

Sucrose

• Inhibits the formation of ice crystals, it gels as it freezes

• Commonly known as table sugar• Maple tree sap is 2% sucrose, the rest is mainly

water

Fructose and Glucose

• Fructose (left) is a simple sugar• Fructose and Glucose are monosaccharides• Glucose is synthesized from glycerol • Fructose and glucose are the two main

constituents of sap from birch trees• Give birch syrup a very distinct flavor

Frogs and Glycerol

Frogs• Both the Gray Tree Frog and the Wood

Frog are able to safely freeze almost solid

• There livers produces high amounts of glycerol– Through osmosis, gylcerol is exchanged for

water in the cells

• Once thawed in the spring the glycerol is synthesized into glucose (energy boost)

The world of freezing Herps• About a dozen species of amphibians and reptiles are known to

tolerate the freezing of their tissues under thermal and temporal conditions that mimic frost exposure in nature (i.e., slow cooling to relatively high subzero temperatures).

• Some species survive freezing at temperatures as low as -6°C and endure freezing episodes lasting more than a month. Fully-frozen animals, in which up to 65-70% of the body fluid has become ice, appear dead - muscle contraction, heartbeat, and breathing have completely ceased.

• The frozen tissues become depleted of oxygen and the cellular energy status declines sharply. Remarkably, these animals arouse after thawing and can soon resume normal physiological and behavioral functions.

• Freeze tolerance is promoted by special physiological adaptations, including an accumulation of certain cryoprotective compounds, a redistribution of bulk water within the body, and an innate tolerance of cells to dehydration.

Distribution of Rana sylvatica

Map obtained from:http://www.exploratorium.edu/frogs/woodfrog/woodfrog_3.html

Wood frogs have adapted to live in the North of the US, Canada and Alaska by freezing solid during winter and thawing early in spring.

Distribution of wood frogs (red)

What is freezing?

• Life is a complex set of electrochemical reactions. The rate at which chemical reactions take place depends on temperature, and usually the lower the temperature the lower the rate. At absolute zero Kelvin (-460°F), the rate is zero. Therefore, lowering the temperature of biological materials such as cells, organs, or entire organisms to absolute zero causes life to stop indefinitely.

• Because we humans are mostly water, however, any journey into the supercool is physically traumatic.

• At about 31°F, the water in our bodies begins to freeze.

• It starts at this temperature (rather than 32°F) because biological water is in the form of a solution, mainly of ions or charged atoms.

• The survival of any cells during freezing depends at minimum on the rate the temperature changes.

• For most organisms, even the slowest cooling results in an assault on their cells that is just too great.

How the freezing begins• Several mechanisms ensure that wood frogs freeze without

supercooling extensively. • First, owing to the highly permeable nature of amphibian skin, ice

surrounding the frog can instantly trigger the freezing of the body fluids.

• Also, the frog’s winter refuge hosts an abundance of ice nucleating agents, such as various mineral particulates, organic acids, and certain microbes, that may cause the frog to freeze. Laboratory experiments suggest that ingestion of these agents promote ice formation in freeze-tolerant frogs.

• In fact, several strains of bacteria expressing potent ice nucleating activity have been cultured from the intestines of winter-collected wood frogs, indicating that such bacteria are retained throughout hibernation (Lee et al. 1995).

• Inoculation by ice or ice-nucleating agents in the winter environment probably is the primary mechanism initiating freezing in amphibians; there is no need for ice-nucleation proteins or other endogenous ice nuclei, as are found in some invertebrates (Costanzo et al. 1999).

Stresses• Extensive freezing solidifies tissues,

arrests vascular circulation, and deprives cells of oxygen.

• Because ice forms only in extracellular spaces, water inside cells is osmotically drawn externally where it joins the growing ice lattice.

• During this process cells may shrink substantially, potentially with damage to membranes and structural support systems.

• Macromolecules and solutes become crowded in a diminishing solvent volume, perhaps with adverse consequences.

• Ice formation within body fluids also poses the threat of mechanical injury by the growing ice lattice, particularly in compact and highly structured tissues and organs.

• Ice fronts may shear and separate tissues, disrupting intercellular communication systems.

• Upon thawing, large pools of dilute fluid form in extracellular spaces. Cell volume, hydroosmotic balance, and energy status must be restored.

• Resulting cell damage is related to the dissolved substances or solutes in biological water, to the cell membrane properties, and to the fact that ice has a very tight crystallographic structure and cannot contain solutes.

• When biological materials freeze, the solution between cells usually freezes first.

• Solutes found in the original solution are ejected and concentrated in the unfrozen space between the ice crystals.

• Cells usually remain unfrozen though supercooled.

Danger of Freezing

Rupturing of cell membranes by intracellular ice crystals

Normal tissue Ice formation inside cells

Images taken from: http://www.oncura.com/German/prostate-cryotherapy.html

Intracellular ice crystals

When ice forms inside cells, the crystals break the membranes damaging the tissue.This is the most common way of freezing, however wood frogs freeze in a different way…

Freezing of Wood Frogs

• When the frog’s body becomes covered by ice the liver is stimulated to produce glucose.

• Their hear rate doubles, aiding in the fast transfer of glucose to all organs.

• The frog’s large cavities are the first to freeze avoiding the formation of intracellular ice crystals which can be deathly.

Images obtained from:www.zoldmagazin.com/belso/fagvottbeka.htmal

http://seattletimes.nwsource.com/html/nationworld/2002118796_frogs14.html

Bacteria and dust particles found in the frog’s large cavities act as nucleators for ice crystal formation. Ice that is growing in these extracellular spaces attracts water out of cells by osmosis.

Images obtained from:www.gsfc.nasa.gov/feature/2004/0116dust.htmlwww.aquat1.ifas.ufl.edu/guide/bacecoli.jpg http://coslabindia.com/fibremodel2.htm

Bacteria Dust Particle

Ice Crystals

• In order to balance out the resulting difference in potential energy between the inside and outside of the cell, water leaves the cell through the cell membrane.

• Inside the cell, this loss of water causes an increase in the ionic concentration and leads to chemical damage. Interestingly, ions, not ice crystals, trigger cell injury during freezing.

• Theoretically, an infinitely fast rate to absolute zero would eliminate this harm. This is not possible, of course, and at higher cooling rates the supercooling of water in cells causes ice to form within those cells, which also brings about damage.

Alternative crystallization process

Ice crystals in wood frogs form outside the cell

Extracellular ice formation Dehydration

Images taken from: http://www.oncura.com/German/prostate-cryotherapy.html

Normal tissue

The extracellular crystals cause dehydration of the cells through osmosis, but the excess of glucose inside the cells protects them from freezing.

Wood Frog Freezing

• Cells in freeze-tolerant wood frogs experience the same mechanism of freezing injury as any other creatures' cells.

• The frogs freeze very slowly to a temperature often several degrees below freezing. This should destroy the frog's cells, yet those cells and the frog as a whole survive.

• By lowering the amount of water that leaves the cell during freezing, the glucose offers protection against the rise in ionic concentration and excessive cell shrinkage, thereby reducing chemical harm.

Glucose C6 H12 06

• Glucose is a monosaccharide. It has 6 carbons (blue) 12 hydrogens (yellow) and 6 oxygens (white).

• It can work as antifreeze, preventing the cells from collapsing or freezing solid.

http://personal.tmlp.com/Jimr57/textbook/chapter2/cs.htm

• The accumulated glucose apparently enhances the survival of cells, tissues, and organs because experimentally administering additional glucose to the frog increases its tolerance to freezing (Costanzo et al. 1993).

• One of the primary functions of glucose is to raise the osmotic pressure of the body fluids, which in turn reduces the amount of ice that forms at any given temperature.

• Glucose transported into cells acts as an osmolyte, decreasing the degree of cell shrinkage during freezing, and also serves as a fermentable fuel that can be metabolized in the absence of oxygen.

Ice crystals and blood flowThe science of cryogenic surgery

This may look like a set of steak knives, but it's actually ice crystals that form in a physiological saline solution.

These images show red blood cells between ice crystals, at different temperatures below freezing. (A) is the lowest subfreezing temperature, (D) the highest. Note how the cells shrink as they become exposed to lower temperatures.

Magnetic Resonance Image of a wood frog in the process of freezing

The dark region is where ice crystals have formed.

The liver is the last organ to freeze; it produces glucose saturating all vital organs and therefore lowering the frog's freezing point.

Image obtained from: http://www.exploratorium.edu/frogs/woodfrog/woodfrog_4.html

Evolutionary limits

• The frogs have evolved to produce just the right composition of cryoprotectants and gross tissue properties that allow them to survive freezing at the temperatures they experience in nature.

• They cannot survive freezing at lower temperatures.

• This is the key attribute of evolution: it solves only the challenge an organism encounters and nothing else. This is the first-ever ultrasound

image of a frozen lesion in a liver. The arrows point to the margin of the frozen lesion, which appears dark because it reflects the ultrasound pressure waves.

Production of Fibrinogen

Another adaptation of wood frogs is the production of “fibrinogen”, a blood-clotting enzyme which helpsstop any bleeding caused by ice.

Recovery• Recovery is remarkably rapid, with basic physiological and

behavioral functions usually returning within several hours of thawing.

• In collaboration with Jack R. Layne, Jr. (Slippery Rock University), our work has shown that recovery dynamics are characterized by sequential restoration of fundamental to progressively more complex functions.

• For example, the heart resumes beating even before ice in the body has completely melted, and pulmonary respiration and blood circulation are restored soon thereafter.

• Contractility in hindlimb muscles returns 1-2 h after thawing, whereas function of the innervating sciatic nerve is restored within approximately 5 h.

• Hindlimb retraction and righting reflexes return several hours later and the frogs usually exhibit normal body postures and coordinated motor functions within 14-24 h.

• Higher order behaviors, such as mating drive and courting behavior, are not restored until at least several days later (Costanzo et al. 1997).

MRI’s showing the thawing process of wood frogs:Dark areas are frozen

Light areas have thawed

• Wood frogs thaw out evenly; if the exterior unfroze before the heart, liver and brain, the limbs would die due to lack of oxygen.

Images obtained from:http://www.exploratorium.edu/frogs/woodfrog/woodfrog_5.html

Eastern Box Turtle

• This species hibernates in shallow excavations in deciduous forests throughout the eastern United States. Insulated from winter's cold only by a thin blanket of leaf litter and snow (when present), these turtles encounter frost in their hibernacula, yet survive the freezing of their tissues

• Our data, not yet fully analyzed, showed that turtles occasionally freeze during winter, with body temperatures falling several degrees below zero. Survival of these freezing episodes was good. However, in January of one winter, turtles were lulled out of their hibernacula by unseasonably high temperatures only to be caught abroad and immobilized by a rapidly approaching Alberta Clipper. Two turtles were killed outright by heavy frost (core body temperatures near –5°C), and another succumbed some months afterwards. Overall, the ability to survive most frost exposures seems to be an important adaptation permitting winter survival in the northern portion of the species’ range.

Painted turtles • Painted turtles (Chrysemys picta)

inhabit freshwater habitats from coast to coast in the northern United States and southern Canada. These turtles generally overwinter underwater, except that the young, which hatch in late summer, commonly hibernate inside the natal nest, only ~10 cm beneath the ground surface. The cold hardiness of painted turtle hatchlings is remarkable, as many emerge from their nests in spring after being exposed to temperatures that may fall to -11°C or below.

• The biochemical and physiological adaptations promoting the extreme cold hardiness seen in some turtles are still incompletely understood. With respect to freeze tolerance, there is no involvement of thermal hysteresis (= antifreeze) proteins or special ice-nucleating proteins; however, the question of cryoprotectants is unresolved.

• Freezing turtles accumulate small quantities of glucose, lactate, and certain amino acids, though it seems doubtful that the concentrations ultimately achieved, which are much lower than those found in frozen frogs, could substantially reduce the body ice content.

• An innate anoxia tolerance likely helps frozen turtles cope with ischemia, though our studies strongly suggest that the cause of freezing mortality is unrelated to oxygen deprivation. Supercooled turtles also accumulate glucose and lactate, but whether this response improves survival is unknown.

• One thing is clear, however: cold acclimatization is crucial to the development of supercooling capacity, inoculation resistance, and freeze tolerance (Costanzo et al. 2000).

Supercooling

• The supercooling capacity of hatchling painted turtles is probably the best of any vertebrate animal, as these turtles - if carefully isolated from the ice nucleating agents naturally found in their nest - may cool to -20°C before spontaneously freezing. Although they may remain unfrozen, hatchlings nevertheless perish at temperatures below -12°C. Still, supercooling provides protection over a remarkably broad range of environmental temperatures, and clearly allows turtles to survive at temperatures much lower than could be tolerated in the frozen state.

The Manitoba red garter snake

• Not only does this little 20 inch snake survive harsh minus forty (- 40) degree cold winter Canadian weather but also travel as much as 50 miles to do so.

• Narcisse, in the Manitoba Interlake region lies between Lake Winnipeg and Lake Manitoba.

• Here you will see more snakes than anywhere else in the world.

• The limestone provides winter dens below the frost line in this special “hibernacula”..

Snake sex

• First out in the springtime, are the male red sided garter snakes.

• Hundreds and even it may seem thousands may emerge all once. The females appear soon after, but singly or in small numbers. The emergence of the female garter snakes is even spread over several weeks.

• As each female garter snake appears at the surface she will be “mobbed” by male suitors. “Mating balls “ are formed , consisting of a single female garter snake intertwined with as many as 100 males

Hibernaculum Frenzy

• These dens are sinkholes in the local limestone rock produced simply when underground caverns have collapsed. The resulting fissures and crevices in the limestone bedrock give the snakes access to depths below the frost line

Manitoba Mania

Conifers, Alcohols and Terpenes

Coniferous Trees• Conifer sap contains alcohols and

terpenes (i.e. alpha- and beta-pinene)

• Very little is understood about the specific chemicals employed by conifers to prevent freezing– It is believed that the alcohols and terpenes

help them survive winter

• Spruce will employ alcohols and terpenes to force the water out of their cells– This prevents cell damage from ice crystals

Alpha- and Beta-Pinene

• Very low melting points (i.e. freezing temperature) down to -60ºC

• Main constituent in conifer sap• In the terpene family of compounds

– Hydrophobic alkenes

Alcohols

• Alcohols depress the freezing point of water• A variety of alcohols are employed by conifers to

survive the winter • Are characterized by an -OH group attached to

alkanes

Mosses and Arachidonic Acid

Mosses

• Many mosses continue photosynthesizing throughout winter

• Some animals (lemmings and reindeer for example) will eat mosses to help them with the low temperatures of winter (behavioral adaptation)– This allows animals to gain the benefit of the

mosses’ natural anitfreeze -arachidonic acid

Arachidonic Acid

• Helps cells keep moving in low temps• Acts to toughen cell membranes to protect

against ice crystals• Is a second messenger to relay signals within a

cell• Makes up 35% of the fatty acids present in moss

Animal heat response

lower critical temperature (LCT) is the turning point at which more heat is lost to the environment than is normally metabolically produced.

The lower lethal temperature (LLT) is the extreme cold temperature where an animal can no longer produce enough heat and dies of hypothermia.

zone of metabolic regulation

Solar collection• Solar radiation plays a large part in

determining not only ambient and body temperatures but animal behavior as well. A wide variety of species have developed methods to reduce the cost of thermoregulation by behaving certain ways: seeking shade, burrowing, panting, gular fluttering, wing flapping when exposed to temperatures above the UCT, entering short bouts of torpor or longer bouts of hibernation, increasing insulation, or migrating.

Most organisms attempt to remain within a favorable range of temperatures. For homeotherms, this is known as the thermal neutral zone (TNZ). This optimal range of ambient temperatures typically lies between 30-42°C Solar Radiation homeothermic or euthermic organisms (Speakman, 2004). Within this range, metabolic rate is minimal. Outside the TNZ, metabolic energy is required to maintain TB within the optimal range. Metabolic activity involving temperature-dependent enzyme activity will not function properly if the animal becomes hypo- or hyperthermic. Thus, thermoregulation is an integral part of an organism’s energy balance.

Emissivity is defined relative to what is known as a black body, a perfect emitter with an emissivity of 1.0. Most animals have an emissivity value within 0.90-0.98, often dependent on surface properties such as fur or skin color. Surfaces either reflect or absorb light to varying degrees contingent on pigment levels and texture that in turn affect emissivity. Dark colors absorb energy within the infrared spectrum, increasing the absorptivity of inanimate objects or organisms compared to light colors which reflect visual wavelengths of solar energy.

Heat Loss

An animal can either increase its heat production or reduce its heat loss to maintain homeothermy.

In a countercurrent heat exchanger, the hot fluid becomes cold, and the cold fluid becomes hot.

A homeothermic animal is in a continual state of dynamic equilibrium between heatproduction and heat loss. The continual adjustment of physiological and behavioralresponses to the changing energy flux in the environment results in short-termtemperature changes in the animal