Cytotoxicity

Cytotoxicity is the quality of being toxic to cells. Cells exposed to a cytotoxic compound can respond in a number of ways. The cells may undergo necrosis, in which they lose membrane integrity and die rapidly as a result of cell lysis; they can stop growing and dividing; or they can activate a genetic program of controlled cell death, termed apoptosis.

There are many ways to measure cytotoxicity, but most involve assessment of cell membrane integrity. Membrane integrity can be evaluated by using vital dyes (such as trypan blue or propidium iodide), by protease biomarkers, with MTT or MTS redox potential assays, or by measuring ATP content. Many of these assays involve colorimetric, fluorescence, or luminescence detection.

With one of the widest ranges of detection technologies on the market today, Molecular Devices can provide the bioanalytical and analytical products that you need to conduct cytotoxicity measurements, from cytometry and high-content screening systems to multi-mode microplate analysis systems to high-throughput cellular screening systems.

Cytotoxicity testing: measuring viable cells, dead cells, and detecting mechanism of cell death.

Testing the effects of compounds on the viability of cells grown in culture is widely used as a predictor of potential toxic effects in whole animals. Among the several alternative assays available, measuring the levels of ATP is the most sensitive, reliable, and convenient method for monitoring active cell metabolism. However, recently developed combinations of methods have made it possible to collect more information from in vitro cytotoxicity assays using standard fluorescence and luminescence plate readers. This chapter describes two assay methods. The first utilizes beetle luciferase for measuring the levels of ATP as a marker of viable cells. The second more recently developed multiplex method relies on selective measurement of three different protease activities as markers for viable, necrotic, and apoptotic cells. Data analysis from the measurement of three marker protease activities from the same sample provides a useful tool to help uncover the mechanism of cell death and can serve as an internal control to help identify assay artifacts.

Atomic Absorption Spectroscopy (AAS)

Atomic absorption spectroscopy (AAS) is a spectroanalytical procedure for the quantitative determination of chemical elements using the absorption of optical radiation (light) by free atoms in the gaseous state.

In analytical chemistry the technique is used for determining the concentration of a particular element (the analyte) in a sample to be analyzed. AAS can be used to determine over 70 different elements in solution or directly in solid samples used in pharmacology, biophysics and toxicology research.

Atomic absorption spectroscopy was first used as an analytical technique, and the underlying principles were established in the second half of the 19th century by Robert Wilhelm Bunsen and Gustav Robert Kirchhoff, both professors at the University of Heidelberg, Germany.

The modern form of AAS was largely developed during the 1950s by a team of Australian chemists. They were led by Sir Alan Walsh at the Commonwealth Scientific and Industrial Research Organisation (CSIRO), Division of Chemical Physics, inMelbourne, Australia.

Atomic absorption spectrometry has many uses in different areas of chemistry such as:

• Clinical analysis: Analyzing metals in biological fluids and tissues such as whole blood, plasma, urine, saliva, brain tissue, liver, muscle tissue, semen
• Pharmaceuticals: In some pharmaceutical manufacturing processes, minute quantities of a catalyst that remain in the final drug product
• Water analysis: Analyzing water for its metal content.

Atomic Absorption Theory

Atomic absorption spectroscopy relies on the Beer-Lambert law to determine the concentration of a particular analyte in a sample. The absorption spectrum and molar absorbance of the desired sample element are known, a known amount of energy is passed through the atomized sample, and by then measuring the quantity of light it is possible to determine the concentration of the element being measured.

Instrumentation

In order to analyze a sample for its atomic constituents, it has to be atomized. The atomizers most commonly used nowadays are flames and electrothermal (graphite tube) atomizers. The atoms should then be irradiated by optical radiation, and the radiation source could be an element-specific line radiation source or a continuum radiation source. The radiation then passes through a monochromatorin order to separate the element-specific radiation from any other radiation emitted by the radiation source, which is finally measured by a detector.

Atomizers

The atomizers most commonly used nowadays are (spectroscopic) flames and electrothermal (graphite tube) atomizers. Other atomizers, such as glow-discharge atomization, hydride atomization, or cold-vapor atomization might be used for special purposes. Here are some atomizers:

• Flame Atomizers
• Electrothermal Atomizers

Specialized atomization techniques

While flame and electrothermal vaporizers are the most common atomization techniques, several other atomization methods are utilized for specialized use.

• Glow-discharge atomization
• Hydride atomization
• Cold vapor atomization

Radiation sources

We have to distinguish between line source AAS (LS AAS) and continuum source AAS (CS AAS). In classical LS AAS, as it has been proposed by Alan Walsh, the high spectral resolution required for AAS measurements is provided by the radiation source itself that emits the spectrum of the analyte in the form of lines that are narrower than the absorption lines.

• Hollow cathode lamps
• Electrodeless discharged lamps
• Deuterium lamps
• Continuum Sources

Spectrometer

As already pointed out above, there is a difference between medium-resolution spectrometers that are used for LS AAS and high-resolution spectrometers that are designed for CS AAS. The spectrometer includes the spectral sorting device (monochromator) and the detector.

• Spectrometers for LS AAS
• Spectrometers for CS AAS

PHYTOCHEMICAL SCREENING

PHYTOCHEMICAL SCREENING

Phytochemical screening - is a process of tracing plant constituents. For example you want to found out if a certain plant contains alkaloids (a plant constituent) then, you will be performing a phytochemical screening procedures for alkaloids (in this case mayer's and Wagner's test). There are general plant constituents that can be performed with a standard test. And these are screening for: Alkaloids, Saponin, Glycosides, Flavonoids, Tannins, Phenols, Anthocyanins and Carbohydrates.

REAGENT

Mayer's reagent- is an alkaloidal precipitating reagent used for the detection of alkaloids in natural products. Mayer’s reagent is freshly prepared by dissolving a mixture of mercuric chloride (1.36 g) and of potassium iodide (5.00 g) in water (100.0 ml). Most alkaloids are precipitated from neutral or slightly acidic solution by Mayer’s reagent (potassiomercuric iodide solution) to give a cream coloured precipitate.

Wagner's reagent- An aqueous solution of iodine and potassium iodide; used for microchemical analysis of alkaloids. Also known as Wagner's solution. Wagner's reagent is used to test for alkaloids. To prepare Wagner's reagent; Dissolve 2g of iodine and 6g of KI in 100ml of water.

INDICATOR

Alkaloids- are also sources of antimicrobial agents.

Flavanoids- are phytochemicals that could act as an antimicrobial agent.

Saponins- May serve sa anti-feedants, and to protect the plan against microbes and fungi.

Carbohydrates- Are most abundant class of organic compounds found in living organism. It is also for protection of bacterial cells as bacterial cell wall polysacharides, made up of meurin.

Anthocyanin- is used as a determinant in the pH and pOH of acids and bases.

Tannins- Which are mainly used in tanning or darkening the color of fabrics.

Phenols- are phytochemicals that could act as an antimicrobial agent.

Glycosides- are antidiarrheal and inhibit release of autocoids and prostaglandins.

PLANTS WITH PHYTOCHEMICAL SCREENING

What is blackberry?

Blackberry is a sprawling shrub growing up to 3 meter in height. The stems and leaves bear thorns, although some thorn free cultivars exist. The dark green compound leaves consist of 3 or 5 leaflets. The flowers resemble small roses and are white or light pink coloured. During late summer and autumn the typical blackberries are formed, which purplish-black coloured.

Parts used

Blackberry leaves, bark and root can be used to make tea. The blackberries (fruits) are also eaten and used in fruit juice and jams.

Phytochemicals

These are typical phytochemicals found in blackberry: blackberry leaves contain tannins, gallotannins, dimeric ellagitannins and flavonoids. blackberries (fruits) are exceptionally rich in phytochemicals mainly, flavonoids, anthocyanins, cyanidin, ellagic acid.

Benefits of blackberry

Blackberry leaves are traditionally used for the treatment of burns and scald. Tea from blackberry (leaves, root and bark) is astringent and used against intestinal disorders such as diarrhoea and dysentery. Chewing of fresh blackberry leaves helps to heal bleeding gums and inflammation of the mouth and troat. 
The anthocyanins in blackberries are responsible for the potential anti-carcinogenic properties. The chemopreventive effects of fresh blackberries is caused by their antioxidant properties. Flavonoids strengthen the blood vessels and capillaries.

Other facts

Blackberry is native to Europe (mainly Mediterranean region) and is cultivated in many countries with moderate climate. There exist various cultivars which produce larger berries. From the blackberry juice a very nice red wine can be produced.

Other names

Bramble, brymbyl, bumble-kite, brameberry, brambleberry.

Bacteria

BACTERIA

Constitute a large domain of prokaryotic microorganism. Typically a few micrometres in length, bacteria have a number of shapes, ranging from spheres to rods and spirals. Bacteria were among the first life forms to appear on Earth, and are present in most of its habitats. Bacteria inhabit soil, water, acidic hot springs, radioactive waste, and the deep portions of Earth's crust. Bacteria also live in symbiotic and parasitic relationships with plants and animals.

DIFFERENT KINDS OF BACTERIA

Coccus

Cocci are round, spherical-shaped bacteria. They can occur as a single bacterium or be arranged in a pair, chain or cluster of bacteria depending on which type of coccus it is. 

Bacillus

Bacilli are rod-shaped bacteria that look like cylinders, arranged singly or in chains. Escherichia coli is a rod-shaped bacteria that normally lives in your intestinal tract without causing disease.

Vibrio

Vibrios are one of three types of bacteria with a spiral-like shape. Vibrios are comma-shaped bacteria that look like a curved rod. They typically live in aquatic environments. Vibrio cholera moves in a darting motion by a single flagellum, a whip like structure.

Spirillum

Spirilla are another subgroup of bacteria with a more rigid spiral shape. 

Spirochete

Spirochetes are long, thin and flexible corkscrew-shaped bacteria. They typically move in a distinctive rotating manner that allows them to be mobile in mucus-lined tissue or where it is viscous.

DESCRIBE THAT EACH TYPE OF BACTERIA CAUSE

Coccus- These bacteria cause many different types of common diseases. Among the more common cocci are Staphylococcus aureus, which appears as a cluster of cocci. Staph aureus can often be found in the nose and on the skin without causing disease, but it is also responsible for causing conditions such as boils, pneumonia, meningitis and toxic shock syndrome. Streptococcus pyogenes occurs as a chain of cocci and causes diseases that originate in the throat or skin, including strep throat and scarlet fever. Neisseria meningitidis occurs in pairs of cocci. This bacteria causes bacterial meningitis, an inflammation of the brain and spinal cord, after entering the body through the nose or throat.

Bacillus- A few strains of E. coli do cause disease that's spread typically by eating or drinking contaminated food or water; a typical symptom is diarrhea. Corynebacterium diphtheriae, another rod-shaped bacterium, infects the respiratory tract and causes diphtheria. Diphtheria causes a thick coating on the back of the nose and throat, making it difficult to swallow or breathe, followed by swelling of the neck and potentially death. Bacillus anthracis is the bacteria that cause anthrax. This rod-shaped bacterium grows in long chains and can infect you through broken skin, ingestion or inhalation.

Vibrio- bacteria that causes cholera. Cholera is an intestinal infection that causes severe diarrhea and dehydration, typically transmitted by drinking contaminated water. It is a very serious disease that can lead to death if not treated promptly.

Spirillum- Causes diarrhea. Campylobacter jejuni is typically acquired in places where sanitation is poor or by eating raw or undercooked poultry. Helicobacter pylori is a similar-shaped bacteria found in your stomach. While some people infected with Helicobacter pylori have no symptoms, it is a common cause of stomach inflammation and ulcers.

Spirochete- Two well-known spirochetes that cause disease in humans are Treponema pallidum and Borrelia burgdorferi. Treponema pallidum causes the sexually transmitted disease syphilis. Infection typically begins as a single sore at the site of infection. Additional lesions or rashes can develop elsewhere on the body if left untreated. Borrelia burgdorferi is transmitted through the bite of an infected tick and causes Lyme disease. Infection with Borrelia burgdorferi causes a typical "bull's-eye" rash. If left untreated, it can affect your heart and nervous system and cause arthritis.

DIFFERENCE BETWEEN GRAM-POSITIVE AND GRAM NEGATIVE BACTERIA

GRAM POSITIVE-  Bacteria are those that are stained dark blue or violet by Gram staining. This is in contrast to Gram-negative bacteria, which cannot retain the crystal violet stain, instead taking up the counterstain (safranin or fuchsine) and appearing red or pink. Gram-positive organisms are able to retain the crystal violet stain because of the high amount of peptidoglycan in the cell wall. Gram-positive cell walls typically lack the outer membrane found in Gram-negative bacteria.

GRAM NEGATIVE- Bacteria are a class of bacteria that do not retain the crystal violet stain used in the Gram staining method of bacterial differentiation, making positive identification possible. The thin peptidoglycan layer of their cell wall is sandwiched between an inner cell membrane and a bacterial outer membrane. In Gram staining, the outer lipid-based membrane of gram-negative bacteria is removed by an alcohol solution. The alcohol also decolorizes the then exposed peptidoglycan layer by dissolving away the previously applied crystal violet. A counterstain (safranin or fuchsine) is then added which recolorizes the bacteria red or pink.

WHAT I LEARNED

* I learned the different kinds of bacteria.

* I learned the disease each type of bacteria cause

* I learned the difference between gram positive and gram negative bacteria.

WHAT I DID

* I search through the internet for information about bacteria and learned much more about it.

I CAN APPLY MY LEARNING TO

* I can apply my learning to research proposal that are needed to be done.

Geological Hazards

1. An earthquake (also known as a quake, tremor or temblor) is the result of a sudden release of energy in the Earth's crust that creates seismic waves. 

2. The three types of earthquakes are:
 a. Tectonic Earthquake - a sudden release of energy
          in the earth's crust or upper mantle, usually
          caused by movement along a fault plane or 
          by volcanic activity and resulting in the
          generation of seismic waves which can be
          destructive

  b. Volcanic Earthquake - is an earthquake induced by the movement of magma.

  c. Landslips Earthquake - caused by landslide. 

3.The causes of earthquakes are:

a. Faulting or shifting of faults. 
b. Volcanic activity/ eruption

4. The focus is where exactly in the crust the earthquake is happening while the epicenter is the point on the surface right above the focus of the earthquake .

5. Fore shocks are earthquakes which precede larger earthquakes in the same location. Aftershocks are smaller earthquakes which occur in the same general area during the days to years following a larger event or "main shock", defined as within 1-2 fault lengths away and during the period of time before the background seismic level has resumed.

6. Earthquakes generate seismic waves which can be detected with a sensitive instrument called a seismograph. 

7. Magnitude measures the energy released at the source of the earthquake. Magnitude is determined from measurements on seismographs. Intensity measures the strength of shaking produced by the earthquake at a certain location. Intensity is determined from effects on people, human structures, and the natural environment.

8. What to Do Before an Earthquake

• Make sure you have a fire extinguisher, first aid kit, a battery-powered radio, a flashlight, and extra batteries at home.
• Learn first aid.
• Learn how to turn off the gas, water, and electricity.

9. What to Do During an Earthquake

• Stay calm! If you're indoors, stay inside. If you're outside, stay outside.
• If you're indoors, stand against a wall near the center of the building, stand in a doorway, or crawl under heavy furniture (a desk or table). Stay away from windows and outside doors.

10. What to Do After an Earthquake

• Check yourself and others for injuries. Provide first aid for anyone who needs it.
• Check water, gas, and electric lines for damage. If any are damaged, shut off the valves. Check for the smell of gas. If you smell it, open all the windows and doors, leave immediately, and report it to the authorities (use someone else's phone).

ITCZ

1. What I learned? 
I learned that The Inter Tropical Convergence Zone (ITCZ), known by sailors as the doldrums, is the area encircling the earth near the equator where the northeast and southeast trade winds come together.

2. What I did is to?
I searched more about this topic.

3. I can apply my learning to?
I can apply my learning by sharing knowledge to other about this topic.

4. I need to learn more about?
I need to learn more things about this topic.

Tropical Depression

1. What I learned?
A tropical depression is an organized system of clouds and thunderstorms with a defined, closed surface circulation and maximum sustained winds of less than 34 knots (39 mph).

2. What I did is to?
I searched more about it to know more things about tropical depression.

3. I can apply my learning to?
I can apply my learning when there is a disaster.

4. I need to learn more about?
I need to learn more about typhoon, Tropical depression, Tropical cyclone and many more.