NORTHEASTERN £49 COLLEGE som setae ous eo CHAPTER V FORENSIC CHEMISTRY 4. Title: THE COMPOSITION OF SEMEN 2. Overviewlintroduction This module focuses on the composition of semen. It specifically provides students the definition, importance and processes in the examination of. It also includes the identification of semen and the ‘examination of semen and seminal stain. Experiment activity is provided at the end of the chapter. 3. Leaming OutcomelObjective At the end of the lesson, the students should be able to: Define Semen Explain the seminal fd Give the process in the examination for Semen Explain cases wherein ejaculation has no spermatozoa Identity where Semen can be found Explain the examination of Semen & Seminal stain onkene 4, Learning ContentTopic Introduction The Seminal Fluid Examination for Semen Cases wherein Ejaculation has No Spermatozoa The Examination of Semen and Seminal Stain The Microscopic Examination of Semen and Seminal Stains Ooeene 5. Teaching and Leaming Activities A. INTRODUCTION ‘The semen refers to the fluid produced by male sex organ. It is usually white to yellowish in colo, ‘consisting of two parts: the seminal plasma or fluid, and the spermatozoa or sperm cells. There are usually 70, (000, 000 to 150, 000, 000 sperm calls per miliiter of semen. The sperm cells, as seen under a microscope, ‘consist of head, a neck and a tall approximately ten times as long 2s the head. During detertoration, the bacteria altack first the tall making identification difficult The seminal fluid contains certain substances called flavins, \Which help give a yellowish color to semen and cause it to flourish under uitra-vioet lights. B, THE SEMINAL FLUID in cases of rape and other sexual crimes, the seminal fluid left by the afackeron the victim's body or on Clothing or furnture presents another powerul source of evidence. Since the sperm inthe fud remain alive foronly a relatively short period, the condition of the sample can give reasonably accurate indications of the time of the altack. In the case of secretors, such as sample contains information of blood group and the presence or ‘absence of other enzymes and proteins that can help to concentrate the search for potential suspects nor rather perpetrators, ‘Sample of seminal stains or fluid are isolated using tests similar to those used to reveal the presence of bloodstains. These tests are particularly useful where attempts have been made by the criminal to remove or wash away incriminating traces. The preceding discussion are intended to explain what these tests are. C. EXAMINATION FOR SEMEN a. WET SPECIMEN b. Normally, suspected semen may be found undried having the alkaline odor characteristics for seminal ‘lud. In this condition, examination is relatively simple, A drop of uid (of semen) is placed on a glass slide, then a drop of distiled water is added. A coverslip is placed over the preparation, The specimen is examined under a high power microscope to determine the presence of sperm cells. DRIED SPECIMEN Examination most offen required is done on dry stan. Physical and chemical tests are used 2s preliminary tests. Confirmation is done by microscopic examination. 1. PHYSICAL EXAMINATION: A general visual examination for grayish-white or yellowish stain is, first made. When dry, semen imparts a starchy stiffness to cloths. When inspected under ultraviolet ight, seminal stain fuoresces. However, this fuorescence isnot specif for semen ‘only but may be observed from other materials or stains. 2. CHEMICAL EXAMINATION: There are three chemical tests that can be used for seminal stains. They are the folowing: a. Florence Test b. Barberio's test; and . Acid Phosphatase Test The first two are based on the formation of characteristics crystals that are observed under the microscope. Acid phosphatase is an enzyme found in both animal and plant cells, but in large concentration in human semen. 3. MICROSCOPIC EXAMINATION: The only specific test for semen is the identification of sperm cell under a microscope. There are many factors which may affect the detection of sperm cells making this method diffcull. Some of these are nature of cloth in which the suspected stain is found, age of stain, condition to which stain was ‘exposed and handling the specimen The presence of sperm cells proves that the stain is semen. However, with the absence of sperm cells, it cannot be concluded that the stain is not of seminal origin. These are some conditions which may lead to non detection of semen, like ASPERMIA, a disorder produced by male organ which produces semen without sperm cells, and OLIGOSPERMIA, a semen with a very few count of sperm cells. D. CASES WHEREIN EJACULATION HAS NO SPERMATOZOA 1. Males suffering from aspermia ‘Aspermia a condition wherein males have no spermatozoa at all in their seminal fuid2. Males suffering from oligospermia Oligospermia- a condition whereby males have abnormally low sperm counts or with few spermatozoa These two diseases can be taken from excessive sexual intercourse. Those suffering from chronic epididynitis and ither testicular diseases. Also taken from chronic venereal diseases, WHERE SEMEN CAN BE FOUND 1. As Fresh a. Vaginal contents ofthe vietim b. Rectal contents ofthe vitim 2. As Wet or Dried Condition a. Hair b. Skin around genitals 3. As Dry Stains a. Underclothing b. Bed Clothing E. THE EXAMINATION OF SEMEN AND SEMINAL STAIN ‘There are four examinations for seminal stains or seminal fluid in the form of stains namely: ‘A. Physical Examination C. Microscopic Examination B. Chemical Examination D. Biological Examination b.1. Florence test b.2. Barberio's test b.3. Acid Phosphatase test 4, THE PHYSICAL EXAMINATION OF SEMINAL STAIN a, Semen when dry gives stif, starchy feeling to the cloth and produces slight deepening of the color with the disappearance of odor. Stiffness disappears if specimen is not property dried in open air. Presence of moisture, bacteria will act on the protein constituent or semen, digest the died protein thus destroy is stifiness. Also the bacteria will remove the albuminous matter and also disintegrate the spermatozoa b. Seminal stain exhibits bluish fluorescence under the ultraviolet light. Ultraviolet light is used to locate invisible seminal stain on cloth. It gives bluish fluorescence provided the cloth is clean and not dark-colored. Bluish fluorescence is not specific for seminal stains and may be seen in some other albuminous materials c. Grayish white, sometimes yellowish stain which is typical of seminal fluid Have appearance or outline of contour map @. May have reddish tint in case of old man 2. THE CHEMICAL EXAMINATION OF SEMINAL STAINS a. Florence Test- thus is known after the name of Dr. Florence of Lysons, who first introduced it Reagent! Chemicals:Florence reagent (1.65 gram potassium iodide and 2.5 grams iodine in 30 co of water) Procedure: 1. Cuta portion of the stain and divide into small bits then soak in saline solution. 2. Transfer into a slide, tease and evaporate the fui. 3, Add a drop of Florence reagent and cover with cover slip. 4, Examine under microscope. \Visible/ Positive Result: Crystals of choline periodide, which ere dark brown, rhombic or needle shaped that occur singly or in crass or even grouped in clusters. It resembles haemin crystals in shape, size and color. Negative reaction maybe due to absence of seminal fluid or spermatic fluid may have not reacted with the reagent due to the very low choline content because of over dilution. Florence testis only preliminary, presence of spermatozoa confirms the presence of seminal stain Limitation of Florence Test: 4, Clothes with seminal stains are not dried thoroughly so choline perioside is decomposed completely, so result is negative. 2. If stan is wet and mixed with blood ‘fier 24 hours itis negative due to rapid decomposition but still spermatozoa can be detected, 4, Even after a long period (2 % years) it will give positive result with Florence test provided thoroughly dried and preserved and if free from blood and other albuminous substance. Ifthe seminal stain contains too much albumen as it is mixed with blood, the albumen interferes to ‘some extent in the test by reacting with so much of the iodine as to leave too litle for the production of Florence's crystals. b, Barberio’s Test Reagent Chemical Saturated aqueous or alcoholic solution of pric acid Procedure: 1. Soak a piece of stained material in 2 2.5% solution of trichloroacetic acid for one hour in a test tube 2. Centrifuge the test tube. 3. Get the clear liquid part and add to an equal amount of a saturated aqueous or alooholic solution of picrc acid on a glass sii. 4, Observe under a microscope. Positive Results: Crystal that are slender yellow tinted, rhomboid needles with obtuse angle or appear 2 ovoid crystals. These crystals are made of specimen picrate.Note: Barberio's test is almost specific for human semen. Seminal stains as old as six years are said to respond to this test. This test is carried out with fresh, dried or dissolved semen. ¢. Acid Phosphatase Test- This testis the best way to locate and at the same time characterize a seminal stain. It has replaced the Florence test in reliability and was shown to be specific for human and higher apes. The test is based fundamentally upon the extraordinarily high acid phosphatase content of human male ejaculate. Phosphatase is the enzyme present in semen. Reagent: Na-@-naphtilphosphate and Fast Blue B Dye Procedure: 41. Moisten with water a piece of filter paper. 2. Swab the stained area with the fiter paper. 3. The acid phosphatase will be transferred tothe fter paper. Add a drop o two of sodium alpha- naphtylphosphate and Fast Blue B dye. Positive result: Purple color. Purple colors indicative of acid phosphatase. Alternative Acid- phosphatase Test Procedure: Reagent: 23 grams of sodium chloride, 0.55 ml of glacial acetic acid, 2 grams of sodium acetate trihydrate in 90 mi water, a suspension of 30 mg of anthraquinone- 1 diazonium chloride and 50 grams of calcium: 1- napthyl phosphate in 1 ml of 1% aerosol Procedure: 41, Treat the stained area in a water bath a pH 5 containing alpha- naphthyl phosphate as a substance and anthraquinone- 1- diazonium chloride. 2, Add the above reagents. Positive result: Orange- red pigment Principle of the Test Aiphanaphtol by the acid phosphatase combines with the diazonium salt to form the color, The reaction takes place for 30 seconds on fresh stains. Limitation ofthe Test: Blood lengthens the time but does not interfere. F, THE MICROSCOPIC EXAMINATION OF SEMEN AND SEMINAL STAINS The chief purpose of microscopic examination is to determine the presence of spermatozoa. The identification of spermatozoa is at present the only specific test for semen.Procedure: Determination of spermatozoa in fresh semen is relatively easier than in stains. Transfer a drop of specimen to a glass slide. ‘Add a drop of water or saline solution and cover with coverslip, Examine under the high power objective. Observe for the presence of spermatozoa. Determination of spermatozoa in seminal stain 1. A piece of material is teased on a slide in a drop of water. 2. Allow the smear to dry and then stain with Loffie’s methylene blue for a minute, wash with water, dry and examine under the microscope. Limitation ofthe Microscopic Test 1. Absence of sperm does not prove that the stain have not been produced by human semen. 2. Elements which may obstruct detection of spermatozoa: a, Nature of fabric b. Age of stain . Condition to which stain was exposed before reaching the laboratory dd. Handling the specimen 3. Some medical jurists believe that there can be no semen without the presence of spermatozoa, but not true in case of aspermia or oligospermia, 6. Flexible Teaching Learning Modality (FTLM) adapted © Module + Google classroom « activities exercises 7. Assessment Task Perform Experiment No. 6