|Year : 2015 | Volume
| Issue : 1 | Page : 24-29
Detection of alcohol in saliva for blood alcohol concentration using alcohol saliva strip test: A forensic aid
Thokala Madhusudhana Rao1, Dorankula Shyam Prasad Reddy1, Pratibha Ramani2, Priya Premkumar2, Natesan Anuja2, Herald J Sherlin2
1 Department of Oral and Maxillofacial Pathology, Kamineni Institute of Dental Sciences, Narketpally, Nalgonda (District), Andhra Pradesh, India
2 Department of Oral and Maxillofacial Pathology, Saveetha Dental College, Saveetha University, Chennai, Tamil Nadu, India
|Date of Web Publication||16-Mar-2015|
Dr. Dorankula Shyam Prasad Reddy
Department of Oral and Maxillofacial Pathology, Kamineni Institute of Dental Sciences, Narketpally, Nalgonda (District), Andhra Pradesh
Source of Support: None, Conflict of Interest: None
Context: Alcohol is a factor in many categories of injury. Alcohol intoxication is frequently associated with injuries from falls, fires, drowning, overdoses, physical and sexual abusements, occupational accidents, traffic accidents and domestic violence. In many instances, for forensic purpose, it may be necessary to establish whether the patients/subjects have consumed alcohol that would have been the reason for the injury/accidents. Combining rapidity and reliability, alcohol saliva strip test (AST) has been put forward for the detection of alcohol in saliva for blood alcohol concentration (BAC). In the present study, we have determined BAC by using AST.
Aims and Objectives: The main objective of this study was to detect alcohol in saliva for BAC in alcoholics by using AST.
Materials and Methods: Two socio-economic groups were selected for the present study where Group A consisted of 40 subjects from the local bar and Group B consisted of 40 subjects from an organized party. The subjects were selected randomly at the local bar and at the organized party who have consumed different forms of alcohol. ALCO-SCREEN 02 plastic strip with a reactive pad was used for the detection of presence of alcohol in saliva.
Results: In the present study, 85% of subjects from Group A, i.e., at the local bar, demonstrated positive results of variable intensity with AST when compared to the subjects from Group B in the organized party which was only about 25%.
Conclusion: The present study showed that AST, performed by using ALCO-SCREEN 02 plastic strip with a reactive pad, can detect the presence of 0.02% BAC or more that can be helpful for various purposes such as forensic, workplace, medical and research settings. The study also showed that amount, time period, concentration and quality of the alcohol intake can influence the BAC, which can be a contributory factor for many accidents, injuries and medical conditions.
Keywords: Alcohol, alcohol saliva strip test, blood alcohol concentration, forensic aid, saliva
|How to cite this article:|
Rao TM, Prasad Reddy DS, Ramani P, Premkumar P, Anuja N, Sherlin HJ. Detection of alcohol in saliva for blood alcohol concentration using alcohol saliva strip test: A forensic aid. J NTR Univ Health Sci 2015;4:24-9
|How to cite this URL:|
Rao TM, Prasad Reddy DS, Ramani P, Premkumar P, Anuja N, Sherlin HJ. Detection of alcohol in saliva for blood alcohol concentration using alcohol saliva strip test: A forensic aid. J NTR Univ Health Sci [serial online] 2015 [cited 2019 Dec 9];4:24-9. Available from: http://www.jdrntruhs.org/text.asp?2015/4/1/24/153310
| Introduction|| |
In these days, alcohol-attributable injuries and violence are of growing concern as alcohol is a factor in many categories of injury.  Each year it is responsible for about 2.3 million premature deaths world-wide. Injuries - both unintentional and intentional - account for more than a third of the burden of disease attributable to alcohol consumption. These include injuries from road traffic crashes, burns, poisoning, falls and drowning as well as violence against oneself or others. The impact of alcohol-related injuries affects not only those who are intoxicated at the time of injury occurrence, but also those who fall victim to their behavior. These include the pedestrian or cyclist knocked over by a drunk driver or woman or children beaten by a drunken husband or father. , Alcohol consumption has been steadily increasing in developing countries like India and decreasing in developed countries since the 1980's.  In India, alcohol consumption is thought to represent a higher socio-economic status. At a snapshot, the scenario of alcohol consumers in India stands at >62.5 million during 1970's that has increased by 106.7% over the 15-year period from 1970 to 1996 and continues to increase at the same pace to the present. ,,
About 80% of alcohol consumption is in the form of hard liquor or distilled spirits showing that the majority drink beverages with a high concentration of alcohol. Branded liquor accounts for about 40% of alcohol consumption, while the rest is in the form of country liquor. People drink at an earlier age than previously. The mean age of initiation of alcohol use has decreased from 23.36 years in 1950-1960 to 18.45 years in 2010. Changing social norms, urbanization, increased availability, high intensity mass marketing and relaxation of overseas trade rules along with poor level of awareness related to alcohol has contributed to increased alcohol use. This alcohol use was associated with 94% of incidents of self-harm, 54% of non-specific, 47% of collapses, 50% of assaults and 50% of patients admitted to hospital. ,, In many instances, for investigation purpose, it may be necessary to establish whether the patients/subjects have consumed alcohol that would have been the reason for the injury/accidents.
The blood alcohol concentration (BAC) is a direct measure of the alcohol level for various purposes such as forensic, workplace, medical and research settings; and several new technologies have been developed for estimating the same. The most preferable method for quantitative measurement of alcohol is gas chromatography for whole blood. However, it is time consuming, expensive and requires skills in laboratory techniques. Reliable, valid and non-invasive methods to quantitatively estimate BAC are important. Until date, non-invasive methods for quantitatively estimating BAC have primarily used breath testing/breathe meters. Although a breath analyzer provides a rapid result, it requires calibration on a regular basis and patient cooperation which may be difficult in combative or comatose patients.  Saliva alcohol saliva strip test (AST) provides a very simple, fast and reliable means for quantitative onsite alcohol detection. Although the precision of the AST is not equivalent to serum determination, it can provide an accurate estimate of BAC in most trauma patients. AST correlates well with serum BAC.  Combining rapidity and reliability, AST using ALCO-SCREEN 02 plastic strip with a reactive pad, has been put forward for the determination of BAC by detecting alcohol in saliva. In the present study, we have determined BAC by using AST. This study evaluated a new device/technique for estimating BAC through saliva that is unique in providing on-the-spot, quantitative results which can be helpful in forensic investigations as an aid.
| Materials and Methods|| |
The present study has been reviewed by the institutional ethical committee and has therefore been performed in accordance with the ethical standards laid down in the 1965 Declaration of Helsinki. The present study was a prospective study measuring BAC by using AST. The procedure for AST was explained to patients and verbal consent was obtained. For patients who were unable to provide consent before the test, verbal consent was obtained once the patient was deemed to be alert and oriented. Two socio-economic groups were selected for the present study where Group A consisted of 40 subjects from the local bar and Group B consisted of 40 subjects from organized party. The subjects were selected randomly at the local bar and at the organized party who have consumed different forms of alcohol and all the subjects ranged from 25 to 40 years of age.
ALCO-SCREEN 02 plastic strip with a reactive pad was used for the detection of presence of 0.02% BAC or more than was imported from Chematics, North Webster, IN, USA. The United States Department of Transportation has established a BAC of 0.02% (0.02 g/dL) as the cut-off level at which an individual is considered positive for the presence of alcohol. ALCO-SCREEN 02 plastic strip contains tetramethylbenzidine (TMB) 0.12 mg, alcohol oxidase (ALOx) 0.5 IU, peroxidase 0.35 IU and proteins 0.15 mg. 
Principle of the procedure
The AST is based on the high specify of ALOx for ethyl alcohol in the presence of peroxidase and enzyme substrate such as TMB as shown in the following:
The distinct color on reactive pad could be observed in <20 s after the tip was contacted with saliva samples with the ethyl alcohol concentration >0.02%. It should be pointed out that other alcohols such as methyl, propyl and allyl alcohol would develop the similar color on the reactive pad. However, these alcohols are not normally present in saliva.
All the subjects were abstained from any intake of coffee, soft drinks, tobacco products for 15 min prior to testing. The ALCO-SCREEN 02 plastic strip with a reactive pad was checked for light cream color in the pads and then the pad was saturated with the saliva of the subjects either by applying directly or from a sputum cup [Figure 1]a. The results on the pad were observed after 2 min. Results after more than 2 min may not be accurate and hence not considered. 
|Figure 1: (a) Demonstration of alcohol saliva strip test. (b) Negative control showing no color change in the reactive pad. (c) Positive control showing color change in the reactive pad|
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Interpretation of results
Almost no color changes by comparing with the background [Figure 1]b. The negative result indicated that the BAC was less than 0.02%.
A distinct color developed all over the pad [Figure 1]c. The positive result indicated that the BAC was 0.02% or higher.
The test was considered as invalid if only the edge of the reactive pad turned color that might be ascribed due to insufficient sampling. The subject was re-tested.
Data obtained were subjected for statistical analysis especially for Pearson Chi-square.
| Results|| |
In the present study, 34 out of 40 subjects (85%) from Group A, i.e., at the local bar, demonstrated positive results of variable intensity with AST while six subjects (15%) tested negative with AST [Table 1], [Figure 2]. The results indicate that P value was found to be statistically significant (P = 0.000) [Table 2]. In Group B, i.e., at the organized party, 10 out of 40 subjects (25%) demonstrated positive results of variable intensity with AST while 30 subjects (75%) tested negative with AST [Table 3], [Figure 3]. The results indicate that P value was found to be statistically significant (P = 0.000) [Table 4] (P < 0.05 was considered as significant).
|Figure 2: Subjects from Group A, i.e., at the local bar, demonstrating positive results of variable intensity with alcohol saliva strip test|
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|Figure 3: Subjects from Group B, i.e., at the organizing party, demonstrating negative and positive results of variable intensity with alcohol saliva strip test|
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|Table 1: 85% Subjects from Group A I.E at The Local Bar, Demonstrate Positive Results with Ast|
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|Table 3: 25% Subjects from Group B I.E at the Organized Party, Demonstrate Positive Results with Ast|
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| Discussion|| |
In many instances, for forensic purposes, it may be necessary to establish whether the patients/subjects have consumed alcohol that would have been the reason for the assault or injury/accidents. Several new technologies such as gas chromatography, breath testing/breathe meters, urine analysis, etc., have been developed for estimating the BAC. , BAC reflects the amount of alcohol in the body which in turn depends on many factors such as food, type and the quantity of beverage, weight, sex and rate of elimination of alcohol.  To determine BAC the above mentioned methods are time consuming, expensive, requires skills in laboratory techniques. Hence, the present study was undertaken in search of a reliable, valid and non-invasive method to quantitatively estimate BAC. Combining rapidity with reliability, the present study was undertaken in search of a technique that measure BAC through saliva sample.
The present study used AST, using ALCO-SCREEN 02 plastic strip with a reactive pad, for the estimation of BAC in two socio-economic Groups, A (local bar) and B (organized party). The results of the present study showed that it is possible to detect BAC of 0.02% or higher with a fair degree of accuracy. The results also showed that P value was found to be statistically significant (P = 0.000) in each group respectively. In the present study, 34 out of 40 subjects (85%) from Group A and 10 out of 40 subjects (25%) in Group B, demonstrated positive results of variable intensity with AST. The positive result is due to consumption of drinks containing high alcohol concentration (country liquors and Indian made foreign liquors containing >42% of alcohol), whereas six subjects in Group A and 30 subjects in Group B showed a negative result because they consumed drinks containing lesser alcohol concentration (beer containing 5% of alcohol). The difference of results in both study groups can be related to the concentration, food, time and quality of the alcohol intake.
Alcohol is absorbed from the stomach and small intestine by diffusion. Most absorption occurs from the small intestine due to its large surface area and rich blood supply. The rate of absorption varies with the emptying time of the stomach. In general, the higher the alcohol concentration of beverage, faster rate of absorption. However, above a certain concentration, the rate of absorption may decrease due to the delayed passage of alcohol from the stomach into the small intestine. The maximum absorption rate is obtained with the consumption of an alcoholic beverage containing approximately 20-45% (by volume or v/v) alcohol solution on an empty stomach which was noted in Group A. The absorption rate may be less when alcohol is consumed with food or when high fluid volume/low alcohol content beverages, such as beer, are consumed which was noted in Group B. ,,,
The total time taken for consumption of alcohol can also influence the distribution of alcohol in blood and saliva. Alcoholic beverages consumed over a long period of time may have minimal absorption. The same can be attributed to the two Groups, A and B, in the present study. Most of the subjects in Group A had tossed down the alcohol almost in a single sip or within a short time period. This sudden spurt of alcohol cannot be metabolized completely and hence, reaches the circulation at a rapid pace thereby attaining maximum concentrations in the blood. Whereas in Group B most of the subjects consumed alcohol (beer) over a long period with small sips along with food. This allows slow absorption rate, ample time for metabolism of alcohol by liver, thereby attaining limited concentrations in the blood. ,
The quality of the drink also plays a significant role in detection of alcohol in saliva. High quality liquors contain refined alcohol along with few additives which prolong the intestinal absorption of alcohol and thereby delay its entry into the blood stream. On the other hand, country liquors have higher amounts of raw alcohol without additives, which readily enters the circulation. This reason could be also accredited to positive results in Group A where most of the subjects consumed country liquors, whereas Group B subjects consumed high quality liquors containing refined alcohol that has led to negative result. ,
The results of our study were in concordance to other studies which were performed to investigative BAC levels using AST. Phair et al. determined BACs by ALCO-SCREEN salivary reagent strip in a series of patients who presented with suspected alcohol intoxication and the results were found to be significant (P < 0.0001). They recommended reagent strips for rapid measurement of BAC.  Bates et al. correlated BACs by alcohol sensitive saliva strips with breath estimates of BAC and found close estimates of BAC with that of saliva strips and breathalyzer; recommending alcohol saliva strips for BAC. 
The present study recommends AST using ALCO-SCREEN 02 plastic strip with a reactive pad for the determination of BAC of 0.02% or higher through saliva that is unique in providing on-the-spot, quantitative results. The AST has few limitations, such as: The AST is designed for use with human saliva only; a positive result indicates only the presence of alcohol and does not indicate or measure intoxication, and there is a possibility that technical or procedural errors, as well other substances in certain foods and medicines (such as strong oxidizers, ascorbic acid, tannic acid, polyphenolic compounds, mercaptans, uric acid, bilirubin and oxalic acid) may interfere with the test and cause false results. However, still the AST has good reliability and validity for the noninvasive, quantitative estimation of BAC and has an edge over the other methods. The advantages of this method are as follows:
- The AST results are not influenced by the presence of blood in the oral cavity,
- The non-invasive nature of AST minimizes the risk of needle stick injuries for staff and multiple needle punctures for patients,
- AST provides a determination of the BAC within 5 min and
- It could also be used in determining postmortem saliva ethanol levels. Last but not least, because of the relatively low cost of the AST the saliva test could be a cost-effective alternative in public health settings where mildly to moderately intoxicated persons are encountered. ,
| Conclusion|| |
The present study suggest that AST using ALCO-SCREEN 02 plastic strip with a reactive pad is a practical, rapid and feasible method to measure BAC (through saliva) of 0.02% or higher in all kinds of subjects including those in unconscious state. As AST using ALCO-SCREEN 02 plastic strip with a reactive pad provided rapid, low cost and easy to interpret measurements, it should be considered as an option for determining BAC in proximity to injury events at the trauma bay and also in postmortem cases for forensic purposes as an aid. This method can also be helpful for screening of individuals for alcohol consumption for the identification of those who might be at risk due to alcohol use, and may serve as a deterrent against inappropriate alcohol consumption.
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[Figure 1], [Figure 2], [Figure 3]
[Table 1], [Table 2], [Table 3], [Table 4]