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\def\citeauthor{~Pallepati~Kishor,
              ~Pallepati~Kavitha,
              ~Siva~Kumar~Peta}
\def\citetitle{Development and Standardization of a Polyherbal Anti Urolithiatic Suspension}
\def\copyrightstmt{This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial- ShareAlike 4.0 License, which allows others to remix, tweak, and build upon the work non-commercially, as long as the author is credited and the new creations are licensed under the identical terms.}
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\def\RunningAuthor{Pallepati Kishor et al.}
\firstPage{68}
\articleType{Original Article}
\receivedDate{15 Feb 2022}
\acceptedDate{04 Mar 2022}
\revisedDate{03 Mar 2022}
\journalVolume{2}
\journalIssue{2}
\journalDoi{fjphs.v2i2.235}
\copyrightYear{2022}
\eISSN{2583-116X}
\journalDoi{fjphs.v2i2.235}
\def\authorCount{3}
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\def\journalTitle{Future Journal of Pharmaceuticals and Health Sciences}

\title{Development and Standardization of a Polyherbal Anti Urolithiatic Suspension}
\author{Pallepati~Kishor\textsuperscript{*}\textsuperscript{1}\textsuperscript{\href{https://orcid.org/0000-0002-0707-4614}{\XeTeXLinkBox{\includegraphics[height=7pt]{orcid-logo-pharmascope.pdf}}}},  
            Pallepati~Kavitha\textsuperscript{2}, 
            Siva~Kumar~Peta\textsuperscript{3}~\\[5pt]{\textsuperscript{1}Department of Pharmacognosy and Phytochemistry, Gokula Krishna College of Pharmacy, Sullurupeta, SPSR Nellore District, Andhra Pradesh, India}~\\{\textsuperscript{2}Department of Pharmaceutics, Gokula Krishna College of Pharmacy, Sullurupeta, SPSR Nellore District, Andhra Pradesh, India}~\\{\textsuperscript{3}Department of Pharmaceutical Chemistry, Gokula Krishna College of pharmacy, Sullurupeta, SPSR Nellore District, Andhra Pradesh, India}}

\begin{abstract}
Renal terrific concentration with respect to stone forming components is mostly recognized to become one of the casual factors such as Calculogenisis. At this work, the preparation and standardization of polyherbal suspension was carried out. Proximate analysis values include percentage of the overall residue, proportion like acetone non-soluble residue, fraction of water soluble ash, percentage of moisture content and percentage of extractive values were analysed for various plants. Preliminary phytochemical analysis of various extracts of the plant revealed the presence of various constituents like glycosides, flavonoids, saponins, steroids etc. Poly-herbal anti-urolithiatic suspension was prepared by combining the prepared extracts of \textit{Tribulus terrestris } L.\textit{, Aerva lanata } L.\textit{, Crataeva religiosa } Hook \& Frost and\textit{ Emblica officinalis } L. with suspension base. The prepared formulation showed good stability and redispersibility.Inside the research project, male mice have been chosen to urolithiasis so because excretory system of male mice starts to resemble that from people and then also existing research show that having the quantity like stone discharge through female mice had been substantially lower. Such research results, thereby stimulate the need for any further research to hold over the antilithiatic effect of the polyherbal suspension to prove that more effective treatment for lithiasis with polyherbal suspension can be achieved. 
\end{abstract}\def\keywordstitle{Keywords}

\begin{keywords}Urolithiasis,\newline Polyherbal Suspension,\newline Antilithiatic Effect,\newline Herbal Treatment,\newline Cystone
\end{keywords}
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\makeatletter{\raggedright{\Large$^\ast$}Corresponding Author\par Name:\ Pallepati~Kishor~\\ Phone:\ +91 9951850662~\\  Email:\ kishorpharmcog@gmail.com
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\section{Introduction}
Urolithiasis is a condition where stones form in the kidneys or the bladder. These stones are made of minerals and salts. Patients with this condition can have a variety of symptoms, depending on how severe it is. Urinary stones are calcifications that form in the kidney, ureter, or bladder \unskip~\cite{1365009:24518658}. They are most common in children and older adults. Urinary stones can be just like a little like a dust particle or even as big as either a tennis ball. Symptoms of urinary stones include blood in your urine, an inability to urinate, pain when urinating, increased thirst and dark urine. If untreated there is a high risk for acute renal failure resulting in kidney problems \unskip~\cite{1365009:24518646}. Urolithiasis is a medical term for bladder, kidney or urethra stones. It can occur in men or women. It mainly affects the elderly but it can happen at any age. The risk factors are obesity, diabetes, recurrent urinary tract infections, constipation and other diseases affecting the urinary tract. Urolithiasis is a bladder disease that causes inflammation of the urinary tract. It also causes your kidneys to fail, which can cause life-threatening complications \unskip~\cite{1365009:24518650}. These stones are made up of uric acid or other materials that collect in your urine. Urolithiasis is caused by an abundance of either uric acid or other compounds in the urine that may be passed out by patients who have certain health conditions such as gout, diabetes mellitus type 2, hyperuricemia syndromes, and various other genetic disorders. This may also occur due to overuse of diuretics or excessive alcohol consumption \unskip~\cite{1365009:24518655}.

Urolithiasis is a condition where the urinary stone is made up of mineral salts. It can occur when urine becomes too concentrated, which leads to a decrease in the fluidity of the urine and causes a buildup of minerals. Alternative treatment options for these patients are mostly herbal remedies, including traditional Indian systems of medicine such as Ayurveda and Unani. Herbal treatments have several advantages - they are inexpensive, they have no side effects and they are not as invasive as other treatments, such as surgery or lithotripsy that may be required to remove stones from the bladder or kidneys \unskip~\cite{1365009:24518643}. Urolithiasis is a medical condition of the urinary tract. Urinary stones are created when urine gets concentrated and the minerals it contains turn into stone-like structures that can clog up and irritate urinary passages or collect in the kidneys. Some people don't know that there are many herbal treatments that can be used for urolithiasis. Herbal remedies can be used to help reduce kidney stones and prevent kidney stone formation. Nature has provided us with a variety of herbs that we can use for treatment such as wintergreen, juniper, wild cherry bark, and garlic \unskip~\cite{1365009:24518656}. These herbs have been shown to help reduce kidney stones or stop them from forming altogether by working on the kidneys to cleanse them and release any fluids before they get stuck in the body. This treatment is one of the most popular and effective remedies used for this disease. It has been clinically proven to be helpful in treating urolithiasis more effectively than any other medication.

Poly-herbal anti-urolithiatic suspension was prepared by combining the prepared extracts of \textit{Tribulus terrestris } L.\textit{, Aerva lanata } L.\textit{, Crataeva religiosa } Hook \& Frost and\textit{ Emblica officinalis } L. with suspension base. The prepared formulation showed good stability and redispersibility. Polyherbal suspension was found to be nontoxic in Wistar albino rats upto 2000mg/kg body weight. Within present research, a kind effort is made to assess the combined effect of \textit{Tribulus terrestris} L. (Zygophyllaceae),\textit{ Aerva lanata } L. (Amaranthaceae) and\textit{ Crataeva religiosa } Hook \& Frost. (Capparidaceae) with in research, male mice have been handpicked of about stimulating urolithiasis since the excretory system of male mice replicates that from people and then also existing research studies have found the quantity like stone discharge through female rats had been substantially less \unskip~\cite{1365009:24518654}. 
    
\section{MATERIALS AND METHODS}
\textbf{Collection of Plant Material}

The fruits of \textit{Tribulus terrestris }L. (Zygophyllaceae), whole plant of\textit{ Aerva lanata } L. (Amaranthaceae), bark of \textit{Crataeva religiosa } Hook \& Frost. (Capparidaceae), and fruits of \textit{Emblica officinalis } L. (Euphorbiaceae) have been gathered from Tirumala hills, Tirupathi.

\textbf{Physico Chemical Parameters}

\textbf{Determination of Moisture Content}

About 2g of such substance has been did weigh inside a watch glass, managed to keep out dried in an oven about as 105\ensuremath{^{o}}C but also hardened for one-time span till the study weight has been acquired. Losing weight over dryers has been mentioned but also variation through load provides a water contents like powder form substances. Combined amount water contents of origin has been mentioned.

\textbf{Determination of Total Ash Value}

Weigh accurately 3g of that whole finely ground substance inside a porous reactor core which has been earlier triggered but also decide to weigh, that whole finely ground substance has bee disperse like a perfect also bottom layer of such reactor core. That whole reactor core has been vaporized till a burning hot substance has been acquired about as 450oc heat then it is available through the cellulose. A reactor core has been allowed to cool but also decided to weigh. That whole methodology has been replicated till the consistent body weight. The proportion of a higher ash has been estimated as regards to a oven drying powder form specimen.

\textbf{Determination of Acid Insoluble Ash Value}

The whole gather necessary residue has been heated as for 25ml like 2n hydrochloride for five minutes. A non-soluble residue has been gathered over residue fewer filtrate paper. A non-soluble residue has been transmitted into another pre-weighed silicon dioxide reactor core, triggered, allowed to cool but also did weigh. That whole methodology has been reproduced until the steady weight has been acquired. The proportion like acid non-soluble residue has been measured as regards to those whole air-dry substances \unskip~\cite{1365009:24518651}.

\textbf{Determination of Water Soluble Ash Value Until the Steady Weight Has Been Acquired. The proportion}

That whole ash content acquired has been heated as for 25ml like chloroform liquid just that 5 minutes. That whole non-soluble matter has been gathered on either a residue fewer filtrate paper but also cleaned as for heated water. It and non-soluble residue has been converted into other pre-weighed silicon dioxide reactor core, triggered just that 15 minutes at such a max temp at 4500C, allowed cooling as well as measured, a methodology has been reproduced to have the steady weight. The load of such non-soluble matter has been subtract as for mass of the total residue, the proportion like highly soluble in water residue has been determined by calculating as regards to reference of aerosol specimen substance.

\textbf{Determination of Extractive Values }

\textbf{Alcohol Soluble Extractive Value}

Approximately 5 gram like granular particles (60-80 mesh) of a crude substance (shade dried) has been measured but also dissolved through potassium iodide bottle as for 100millilitre like 70\% v/v alcohol, for such a soan like 24 hour as for common shake and eventually made to stand for 18 h. Solution was filtered rapidly, taking precaution against loss of alcohol, 25 ml of filtered solution was evaporated to dryness at 105\ensuremath{^{o}}C in a tarred flat bottomed petri dish. The percentage of alcohol soluble extract was determined with reference to the shade dried drug. 

\textbf{Water Soluble Extractive Value}

About 5 g of coarse powder (60-80 mesh) of crude drug (shade dried) was weighed and macerated in iodine flask with 100 ml of chloroform-water, for a duration of 24 h, with frequent shaking and finally allowed to stand for 18 h. Solution was filtered rapidly, 25 ml of filtered solution was evaporated to dryness at 105\ensuremath{^{o}}C in a tarred flat bottomed petri dish. The percentage of water soluble extract was determined with reference to the shade dried drug \unskip~\cite{1365009:24518640}.

\textbf{Phyto Chemical Analysis}

Phyto chemicals have played an important role because simple origin for such formation among several biopharmaceuticals. Secondary chemical constituents are chiefly responsible for the biological activities of plants or drugs. The concentrated extracts were subjected to chemical tests as per the methods \unskip~\cite{1365009:24518642}.

\textbf{Preparation of Extracts}

Fruits of \textit{Tribulus terrestris} L, Bark of \textit{crataeva religiosa} Hook \& Frost., Fruits of \textit{Emblica officinalis} L., have been gathered, cleaned and dried there as ambient temperature. Now since full dryers, those were all powder form as well as did pass throughout a 60 mesh sifter but also kept along tightly sealed vessel. Hardened powder form substances were being used to organize samples. Most of 50g like oven drying powder form tree components seem to be harvested with ethanol (60-80\ensuremath{^{o}}C), petroleum ether (60-80\ensuremath{^{o}}C), 80\% methanol (60-80\ensuremath{^{o}}C), in soxhlet apparatus. Extracts were filtered, the solvent was evaporated and accurate weight of the extracts was taken. Extractive values (in \%) were calculated with reference to air dried drug. Color and consistency of extracts were noted.

Whole plant of \textit{Aerva lanata} L. Has been gathered, cleaned and dried about as ambient temperature. Now since entire dryers, this was powder form as well as managed to pass throughout a 60 mesh sifter but also kept out airtight seal vessel. Hardened powder form substance has been used to make preparations extricate. About 50 g of air dried powdered plant material was macerated in iodine flask with chloroform water for duration of 24 h, with frequent shaking and finally allowed to stand for 18 h. Extract was filtered, the solvent was evaporated and accurate weight of the extract was taken. Extractive value (in \%) was calculated with reference to air dried drug. Colour and consistency of extracts were noted \unskip~\cite{1365009:24518653}.

\textbf{Formulation of Anti Urolithiatic Suspension}

Formulation of poly-herbal anti-urolithiatic suspension was prepared by combining the prepared extracts of\textit{ Tribulus terrestris} L. (Zygophyllaceae)\textit{, Aerva lanata} L. (Amaranthaceae)\textit{, Crataeva religiosa } Hook \& Frost. (Capparidaceae) and\textit{ Emblica officinalis} L. (Euphorbiaceae) With such a best suited rescinding agent and same composition. Sufficient amount like sugar solution has been considered to such a sweep engine as well as triturates this into powder and that it was dispersed out 50ml of liquid [Table~\ref{tw-af3241d9c932}]. A harvests, tween 80 as well as sodium cmc through corresponding qty have been captured and also were created it into homogeneous mixture. Such a mixture was transferred to 50ml like sugar solution. Therefore the emigrants such as sorbitol, flavor enhancer operator but also cryoprotectant have been got to add and was manufactured it into a homogeneous mixture. At last the quantity has been created up to nearly 100ml \unskip~\cite{1365009:24518657}.


\begin{table*}[!htbp]
\caption{\boldmath {The Composition of Poly-Herbal Suspension} }
\label{tw-af3241d9c932}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1847\linewidth-2\tabcolsep}p{\dimexpr.4069\linewidth-2\tabcolsep}p{\dimexpr.4084\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S. No & \cAlignHack Drug & \cAlignHack Quantity (100ml)\\
\tblmidrule 
\cAlignHack 1 &
  \cAlignHack \textit{Tribulus terrestris } &
  \cAlignHack 500mg\\
\cAlignHack 2 &
  \cAlignHack \textit{Crataeva religiosa} &
  \cAlignHack 500mg\\
\cAlignHack 3 &
  \cAlignHack \textit{Aerva lanata } &
  \cAlignHack 2.26g\\
\cAlignHack 4 &
  \cAlignHack \textit{Emblica officinalis } &
  \cAlignHack 2.50g\\
\cAlignHack 5 &
  \cAlignHack Tween 80 &
  \cAlignHack 0.1\%\\
\cAlignHack 6 &
  \cAlignHack Sodium CMC  &
  \cAlignHack 2 g\\
\cAlignHack 7 &
  \cAlignHack Sucrose  &
  \cAlignHack 10 g\\
\cAlignHack 8 &
  \cAlignHack Sorbitol  &
  \cAlignHack 5 g\\
\cAlignHack 9 &
  \cAlignHack Methyl paraben &
  \cAlignHack 0.20\%\\
\cAlignHack 10 &
  \cAlignHack Purified water &
  \cAlignHack q.s\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Experimental Design} }
\label{tw-797a4804d7e5}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.13180000000000001\linewidth-2\tabcolsep}p{\dimexpr.2747999999999999\linewidth-2\tabcolsep}p{\dimexpr.28330000000000005\linewidth-2\tabcolsep}p{\dimexpr.31009999999999994\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S.no & \cAlignHack Experimental Design & \cAlignHack Group & \cAlignHack Treatment\\
\tblmidrule 
\cAlignHack  I.  &
  \cAlignHack  &
  \cAlignHack Normal control rats &
  \cAlignHack Normal healthy rats\\
\cAlignHack  II.  &
  \cAlignHack Control rats &
  \cAlignHack Negative control rats &
  \cAlignHack Urolithic rats\\
\cAlignHack  III.  &
  \cAlignHack  &
  \cAlignHack Positive control rats &
  \cAlignHack Urolithic rats receive standard drug(from 15\ensuremath{^{th}} to 28\ensuremath{^{th}} day)\\
\cAlignHack  IV.  &
  \multicolumn{1}{p{\dimexpr(.2747999999999999\linewidth-2\tabcolsep)}}{\multirow{2}{\linewidth}{\cAlignHack Preventive study of formulation}} &
  \cAlignHack Test group-1 &
  \cAlignHack Normal Rats receive inducing agent + formulation (from 1\ensuremath{^{st}} day) {\textendash} 300mg/kg\\
\cAlignHack  V.  &
   &
  \cAlignHack Test group-2 &
  \cAlignHack Normal Rats receive inducing agent + formulation (from 1\ensuremath{^{st}} day) {\textendash} 600mg/kg\\
\cAlignHack  VI.  &
  \multicolumn{1}{p{\dimexpr(.2747999999999999\linewidth-2\tabcolsep)}}{\multirow{2}{\linewidth}{\cAlignHack Curative study of formulation}} &
  \cAlignHack Test group-3 &
  \cAlignHack Urolithic rats receive formulation (from 15\ensuremath{^{th}} day to 28\ensuremath{^{th}} day) -300mg/kg\\
\cAlignHack VII.  &
   &
  \cAlignHack Test group-4 &
  \cAlignHack Urolithic rats receive formulation (from 15\ensuremath{^{th}} day to 28\ensuremath{^{th}} day) {\textendash} 600mg/kg\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Moisture Content of Powdered Plant Materials} }
\label{tw-1f0d642d5821}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.13499999999999998\linewidth-2\tabcolsep}p{\dimexpr.4253999999999999\linewidth-2\tabcolsep}p{\dimexpr.4396\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S.No & \cAlignHack Plant name & \cAlignHack Moisture content (\% W/W)\\
\tblmidrule 
\cAlignHack 1. &
  \cAlignHack \textit{Tribulus terrestris} L. &
  \cAlignHack 4\%w/w\\
\cAlignHack 2. &
  \cAlignHack \textit{Aerva lanata }L. &
  \cAlignHack 5\%w/w\\
\cAlignHack 3. &
  \cAlignHack \textit{Crataeva religiosa }Hook \& Frost. &
  \cAlignHack 5\%w/w\\
\cAlignHack 4. &
  \cAlignHack \textit{Emblica officinalis} L. &
  \cAlignHack 6\%w/w\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Ash Values of Powdered Plant Materials} }
\label{tw-2a8c83f44440}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.06890000000000001\linewidth-2\tabcolsep}p{\dimexpr.35269999999999996\linewidth-2\tabcolsep}p{\dimexpr.1735\linewidth-2\tabcolsep}p{\dimexpr.2165\linewidth-2\tabcolsep}p{\dimexpr.1884\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S. No & \cAlignHack Name of the Plant & \cAlignHack Total Ash (\%w/w) & \cAlignHack Acid Insoluble Ash (\%w/w) & \cAlignHack Water Soluble Ash (\%w/w)\\
\tblmidrule 
\cAlignHack 1 &
  \cAlignHack \textit{Tribulus terrestris} L. &
  \cAlignHack 15\%w/w &
  \cAlignHack 2\%w/w &
  \cAlignHack 6\%w/w\\
\cAlignHack 2 &
  \cAlignHack \textit{Aerva lanata} L. &
  \cAlignHack 14.22\%w/w &
  \cAlignHack 1.67\%w/w &
  \cAlignHack 8.22\%w/w\\
\cAlignHack 3 &
  \cAlignHack \textit{Crataeva religiosa} Hook \& Frost. &
  \cAlignHack 13\%w/w &
  \cAlignHack 1\%w/w &
  \cAlignHack 6.8\%w/w\\
\cAlignHack 4 &
  \cAlignHack \textit{Emblica officinalis} L. &
  \cAlignHack 5\%w/w &
  \cAlignHack 1.5\%w/w &
  \cAlignHack 3\%w/w\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Extractive Values of Powered Plant Materials} }
\label{tw-a3f602c422d1}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.14\linewidth-2\tabcolsep}p{\dimexpr.3578999999999999\linewidth-2\tabcolsep}p{\dimexpr.2459\linewidth-2\tabcolsep}p{\dimexpr.2562\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S. No. & \cAlignHack Plant name & \multicolumn{2}{p{\dimexpr(.5021\linewidth-2\tabcolsep)}}{\cAlignHack Extractive value \%w/w}\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & \cAlignHack Water soluble & \cAlignHack Alcohol soluble\\
\tblmidrule 
\cAlignHack 1 &
  \cAlignHack \textit{Tribulus terrestris} L. &
  \cAlignHack 7.46 &
  \cAlignHack 14.92\\
\cAlignHack 2 &
  \cAlignHack \textit{Aerva lanata} L. &
  \cAlignHack 22.12 &
  \cAlignHack 15.34\\
\cAlignHack 3 &
  \cAlignHack \textit{Crataeva religiosa} Hook \& Frost. &
  \cAlignHack 1.16 &
  \cAlignHack 3.82\\
\cAlignHack 4 &
  \cAlignHack \textit{Emblica officinalis} L. &
  \cAlignHack 9.5 &
  \cAlignHack 16.5\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Characteristics of the Plant Extracts} }
\label{tw-a9acd22d1261}
\centering 
\begin{threeparttable}

\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.0825\linewidth-2\tabcolsep}p{\dimexpr.34669999999999995\linewidth-2\tabcolsep}p{\dimexpr.13239999999999997\linewidth-2\tabcolsep}p{\dimexpr.1241\linewidth-2\tabcolsep}p{\dimexpr.171\linewidth-2\tabcolsep}p{\dimexpr.14329999999999998\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack S.no & \cAlignHack Plant name & \cAlignHack Solvent & \cAlignHack Colour & \cAlignHack Odour & \cAlignHack Consistency\\
\tblmidrule 
\cAlignHack 1 &
  \cAlignHack \textit{Tribulus terrestris} L. &
  \cAlignHack Ethanol &
  \cAlignHack DG &
  \cAlignHack Characteristic &
  \cAlignHack Solid mass\\
\cAlignHack 2 &
  \cAlignHack \textit{Aerva lanata} L. &
  \cAlignHack Aqueous &
  \cAlignHack DG  &
  \cAlignHack Characteristic &
  \cAlignHack Sticky mass\\
\cAlignHack 3 &
  \cAlignHack \textit{Crataeva religiosa} Hook \& Frost. &
  \cAlignHack Pet ether &
  \cAlignHack DG &
  \cAlignHack Characteristic  &
  \cAlignHack Non greesy\\
\cAlignHack 4 &
  \cAlignHack \textit{Emblica officinalis} L. &
  \cAlignHack Methnol &
  \cAlignHack Black  &
  \cAlignHack Characteristic  &
  \cAlignHack Sticky mass\\
\tblbottomrule 
\end{tabulary}\par 
\begin{tablenotes}\footnotesize 
    
\item{DG - Dark Green, BG -  Black Green, DB - Dark Brown}
\end{tablenotes}
\end{threeparttable}

\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Preliminary Phytochemical Analysis of Various Extracts} }
\label{tw-3505c5be94a4}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1983\linewidth-2\tabcolsep}p{\dimexpr.1603\linewidth-2\tabcolsep}p{\dimexpr.2232\linewidth-2\tabcolsep}p{\dimexpr.21990000000000002\linewidth-2\tabcolsep}p{\dimexpr.1983\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Plant Name & \cAlignHack \textit{T. terrestris} (Ethanol) & \cAlignHack \textit{A. lanata }(Aqueous) & \cAlignHack \textit{C. religiosa }(Petroleum ether) & \cAlignHack \textit{E. officinalis} (Methanol)\\
\tblmidrule 
\cAlignHack Alkaloids &
  \cAlignHack \_ &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack +\\
\cAlignHack Carbohydrates &
  \cAlignHack \_ &
  \cAlignHack \_ &
  \cAlignHack \_ &
  \cAlignHack +\\
\cAlignHack Glycosides &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack \_ &
  \cAlignHack +\\
\cAlignHack Steroids &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack + &
  \cAlignHack \_\\
\cAlignHack Fixed oils and Fats &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack + &
  \cAlignHack \_\\
\cAlignHack Saponins &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack - &
  \cAlignHack \_\\
\cAlignHack Phenolic compounds &
  \cAlignHack \_ &
  \cAlignHack \_ &
  \cAlignHack - &
  \cAlignHack \_\\
\cAlignHack Tannins &
  \cAlignHack \_ &
  \cAlignHack + &
  \cAlignHack - &
  \cAlignHack \_\\
\cAlignHack Proteins &
  \cAlignHack \_ &
  \cAlignHack \_ &
  \cAlignHack - &
  \cAlignHack +\\
\cAlignHack Gums and Mucilages &
  \cAlignHack + &
  \cAlignHack \_ &
  \cAlignHack - &
  \cAlignHack +\\
\cAlignHack Flavanoids &
  \cAlignHack + &
  \cAlignHack + &
  \cAlignHack - &
  \cAlignHack +\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Physical Evaluation Values for Formulated Suspension} }
\label{tw-063b951f7bb0}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1104\linewidth-2\tabcolsep}p{\dimexpr.08430000000000001\linewidth-2\tabcolsep}p{\dimexpr.1518\linewidth-2\tabcolsep}p{\dimexpr.1981\linewidth-2\tabcolsep}p{\dimexpr.1672\linewidth-2\tabcolsep}p{\dimexpr.14120000000000003\linewidth-2\tabcolsep}p{\dimexpr.14700000000000001\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}Stability & pH & Viscosity (Cps) & Density (gm/ml) & Sedimentation volume (ml) & Particle size (\ensuremath{\mu }m) & Redispersi \mbox{}\protect\newline bility\\
\tblmidrule 
Good &
  5.03 &
  52.6 &
  5.04 &
  7ml &
  60.64 &
  Good\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Acute Toxicity Test Studies of Poly Herbal Suspension} }
\label{tw-eb00541d06fc}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.0703\linewidth-2\tabcolsep}p{\dimexpr.0816\linewidth-2\tabcolsep}p{\dimexpr.1988\linewidth-2\tabcolsep}p{\dimexpr.1376\linewidth-2\tabcolsep}p{\dimexpr.1262\linewidth-2\tabcolsep}p{\dimexpr.1055\linewidth-2\tabcolsep}p{\dimexpr.12\linewidth-2\tabcolsep}p{\dimexpr.16\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Sl. No & \cAlignHack Animals & \cAlignHack PHS \mbox{}\protect\newline Dose/kg (body weight) p.o & \multicolumn{2}{p{\dimexpr(.2638\linewidth-2\tabcolsep)}}{\cAlignHack Weight of Rats } & \cAlignHack Signs of Toxicity & \cAlignHack Onset of Toxicity & \cAlignHack Duration of Study\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & \cAlignHack  & \cAlignHack Before Test (gm) & \cAlignHack After Test (gm) & \cAlignHack  & \cAlignHack  & \cAlignHack \\
\tblmidrule 
\cAlignHack 1. &
  \cAlignHack Rat 1 &
  \cAlignHack 2000 mg &
  \cAlignHack 168 &
  \cAlignHack 170 &
  \cAlignHack No signs of Toxicity &
  \cAlignHack Nil &
  \cAlignHack 14 days\\
\cAlignHack 2. &
  \cAlignHack Rat 2 &
  \cAlignHack 2000 mg &
  \cAlignHack 172 &
  \cAlignHack 173 &
  \cAlignHack No signs of Toxicity &
  \cAlignHack Nil &
  \cAlignHack 14 days\\
\cAlignHack 3. &
  \cAlignHack Rat 3 &
  \cAlignHack 2000 mg &
  \cAlignHack 187 &
  \cAlignHack 186 &
  \cAlignHack No signs of Toxicity &
  \cAlignHack Nil &
  \cAlignHack 14 days\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Urinary Excretion of Stone Forming Constituents in Control and Experimental Rats} }
\label{tw-36d0f6a25c6a}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.19999999999999996\linewidth-2\tabcolsep}p{\dimexpr.1884\linewidth-2\tabcolsep}p{\dimexpr.2116\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Groups & \cAlignHack Dose (mg/kg) & \multicolumn{3}{p{\dimexpr(.6116\linewidth-2\tabcolsep)}}{\cAlignHack Urine Parameters}\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & Calcium  \mbox{}\protect\newline (mg/24 hr) & Oxalate \mbox{}\protect\newline (mg/24 hr) & Phosphate (mg/24 hr)\\
\tblmidrule 
\cAlignHack Group-I \mbox{}\protect\newline Normal control &
  \cAlignHack Vehicle &
  1.52\ensuremath{\pm}0.08812 &
  0.35\ensuremath{\pm}0.01493 &
  3.44\ensuremath{\pm}0.05498\\
\cAlignHack Group-II \mbox{}\protect\newline Calculi induced &
  \cAlignHack Vehicle &
  4.47\ensuremath{\pm}0.1441 &
  4.11\ensuremath{\pm}0.05412 &
  7.46\ensuremath{\pm}0.05828\\
\cAlignHack Group-III \mbox{}\protect\newline Cystone treated &
  \cAlignHack 750 &
  1.50\ensuremath{\pm}0.05357 &
  0.48\ensuremath{\pm}0.02982 &
  3.64\ensuremath{\pm}0.1288\\
\cAlignHack Group-IV \mbox{}\protect\newline Curative regimen-1 &
  \cAlignHack 300 &
  1.75\ensuremath{\pm}0.02836 &
  1.26\ensuremath{\pm}0.01745 &
  4.70\ensuremath{\pm}0.1762\\
\cAlignHack Group-V \mbox{}\protect\newline Curative regimen-2 &
  \cAlignHack 600 &
  1.55\ensuremath{\pm}0.03008 &
  0.76\ensuremath{\pm}0.0629 &
  3.94\ensuremath{\pm}0.166\\
\cAlignHack Group-VI \mbox{}\protect\newline Preventive regimen-1 &
  \cAlignHack 300 &
  1.44\ensuremath{\pm}0.0348 &
  0.72\ensuremath{\pm}0.06222 &
  3.97\ensuremath{\pm}0.04139\\
\cAlignHack Group-VII \mbox{}\protect\newline Preventive regimen-2 &
  \cAlignHack 600 &
  1.36\ensuremath{\pm}0.02556 &
  0.52\ensuremath{\pm}0.07306 &
  3.69\ensuremath{\pm}0.07681\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Kidney Retention of Stone Forming Constituents in Control and Experimental Rats} }
\label{tw-b2a49dfd5a12}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1967\linewidth-2\tabcolsep}p{\dimexpr.2033\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Groups & \cAlignHack Dose (mg/kg) & \multicolumn{3}{p{\dimexpr(.60\linewidth-2\tabcolsep)}}{\cAlignHack Kidney parameters}\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & Calcium \mbox{}\protect\newline (mg/g) & Oxalate \mbox{}\protect\newline (mg/g) & Phosphate \mbox{}\protect\newline (mg/g)\\
\tblmidrule 
\cAlignHack Group-I \mbox{}\protect\newline Normal control &
  \cAlignHack Vehicle &
  3.24\ensuremath{\pm}0.02676 &
  1.38\ensuremath{\pm}0.007032 &
  2.33\ensuremath{\pm}0.02777\\
\cAlignHack Group-II \mbox{}\protect\newline Calculi induced &
  \cAlignHack Vehicle &
  4.80\ensuremath{\pm}0.04201 &
  6.74\ensuremath{\pm}0.04738 &
  3.92\ensuremath{\pm}0.05029\\
\cAlignHack Group-III \mbox{}\protect\newline Cystone treated &
  \cAlignHack 750 &
  3.26\ensuremath{\pm}0.02076 &
  1.48\ensuremath{\pm}0.01662 &
  2.32\ensuremath{\pm}0.02553\\
\cAlignHack Group-IV \mbox{}\protect\newline Curative regimen-1 &
  \cAlignHack 300 &
  3.81\ensuremath{\pm}0.05845 &
  1.91\ensuremath{\pm}0.01662 &
  3.43\ensuremath{\pm}0.02459\\
\cAlignHack Group-V \mbox{}\protect\newline Curative regimen-2 &
  \cAlignHack 600 &
  3.44\ensuremath{\pm}0.03194 &
  1.51\ensuremath{\pm}0.009457 &
  2.43\ensuremath{\pm}0.01342\\
\cAlignHack Group-VI \mbox{}\protect\newline Preventive regimen-1 &
  \cAlignHack 300 &
  3.74\ensuremath{\pm}0.02895 &
  1.71\ensuremath{\pm}0.009457 &
  2.70\ensuremath{\pm}0.01978\\
\cAlignHack Group-VII \mbox{}\protect\newline Preventive regimen-2 &
  \cAlignHack 600 &
  3.32\ensuremath{\pm}0.02503 &
  1.40\ensuremath{\pm}0.01327 &
  2.29\ensuremath{\pm}0.01978\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Serum Parameters in Control and Experimental Rats} }
\label{tw-9d2a50d47447}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.20\linewidth-2\tabcolsep}p{\dimexpr.1718\linewidth-2\tabcolsep}p{\dimexpr.2282\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}p{\dimexpr.20\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Groups & \cAlignHack Dose (mg/kg) & \multicolumn{3}{p{\dimexpr(.6282\linewidth-2\tabcolsep)}}{\cAlignHack Serum Parameters}\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & BUN \mbox{}\protect\newline (mg/dl) & Creatinine \mbox{}\protect\newline (mg/dl) & Uric acid \mbox{}\protect\newline (mg/dl)\\
\tblmidrule 
\cAlignHack Group-I \mbox{}\protect\newline Normal control &
  \cAlignHack Vehicle &
  18.45\ensuremath{\pm}0.08888 &
  0.72\ensuremath{\pm}0.0139 &
  0.64\ensuremath{\pm}0.0152\\
\cAlignHack Group-II \mbox{}\protect\newline Calculi induced &
  \cAlignHack Vehicle &
  24.69\ensuremath{\pm}0.1461 &
  1.73\ensuremath{\pm}0.01585 &
  1.42\ensuremath{\pm}0.01956\\
\cAlignHack Group-III \mbox{}\protect\newline Cystone treated &
  \cAlignHack 750 &
  18.81\ensuremath{\pm}0.02837 &
  0.83\ensuremath{\pm}0.01195 &
  0.84\ensuremath{\pm}0.01506\\
\cAlignHack Group-IV \mbox{}\protect\newline Curative regimen-1 &
  \cAlignHack 300 &
  21.19\ensuremath{\pm}0.1717 &
  1.14\ensuremath{\pm}0.02693 &
  1.15\ensuremath{\pm}0.02072\\
\cAlignHack Group-V \mbox{}\protect\newline Curative regimen-2 &
  \cAlignHack 600 &
  19.02\ensuremath{\pm}0.01606 &
  0.97\ensuremath{\pm}0.01118 &
  0.89\ensuremath{\pm}0.01014\\
\cAlignHack Group-VI \mbox{}\protect\newline Preventive regimen-1 &
  \cAlignHack 300 &
  20.44\ensuremath{\pm}0.1352 &
  0.99\ensuremath{\pm}0.03528 &
  0.91\ensuremath{\pm}0.01579\\
\cAlignHack Group-VII \mbox{}\protect\newline Preventive regimen-2 &
  \cAlignHack 600 &
  18.62\ensuremath{\pm}0.06779 &
  0.77\ensuremath{\pm}0.03528 &
  0.81\ensuremath{\pm}0.01579\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\begin{table*}[!htbp]
\caption{\boldmath {Urine Volume, Urine pH, Kidney Weight in Control and Experimental Rats } }
\label{tw-10940ab2f0f0}
\def\arraystretch{1.1}
\ignorespaces 
\centering 
\begin{tabulary}{\linewidth}{p{\dimexpr.1867\linewidth-2\tabcolsep}p{\dimexpr.2067\linewidth-2\tabcolsep}p{\dimexpr.2182\linewidth-2\tabcolsep}p{\dimexpr.16029999999999998\linewidth-2\tabcolsep}p{\dimexpr.2281\linewidth-2\tabcolsep}}
\tbltoprule \rowcolor{kwdboxcolor}\cAlignHack Groups & \cAlignHack Dose (mg/kg) & \multicolumn{3}{p{\dimexpr(.6066\linewidth-2\tabcolsep)}}{\cAlignHack Parameters}\\
\rowcolor{kwdboxcolor}\cAlignHack  & \cAlignHack  & \cAlignHack Urine Volume (ml) & \cAlignHack Urine pH & \cAlignHack Kidney weight (mg)\\
\tblmidrule 
\cAlignHack Group-I \mbox{}\protect\newline Normal control &
  \cAlignHack Vehicle &
  \cAlignHack 14.7\ensuremath{\pm}0.1765 &
  \cAlignHack 6.68\ensuremath{\pm}0.0351 &
  \cAlignHack 0.52\ensuremath{\pm}0.004216\\
\cAlignHack Group-II \mbox{}\protect\newline Calculi induced &
  \cAlignHack Vehicle &
  \cAlignHack 9.88\ensuremath{\pm}0.1407 &
  \cAlignHack 5.76\ensuremath{\pm}0.04118 &
  \cAlignHack 0.68\ensuremath{\pm}0.01647\\
\cAlignHack Group-III \mbox{}\protect\newline Cystone treated &
  \cAlignHack 750 &
  \cAlignHack 18.4\ensuremath{\pm}0.2365 &
  \cAlignHack 6.59\ensuremath{\pm}0.04006 &
  \cAlignHack 0.43\ensuremath{\pm}0.01647\\
\cAlignHack Group-IV \mbox{}\protect\newline Curative regimen-1 &
  \cAlignHack 300 &
  \cAlignHack 13.54\ensuremath{\pm}0.2499 &
  \cAlignHack 6.06\ensuremath{\pm}0.01701 &
  \cAlignHack 0.59\ensuremath{\pm}0.01211\\
\cAlignHack Group-V \mbox{}\protect\newline Curative regimen-2 &
  \cAlignHack 600 &
  \cAlignHack 16.13\ensuremath{\pm}0.1907 &
  \cAlignHack 6.39\ensuremath{\pm}0.02431 &
  \cAlignHack 0.47\ensuremath{\pm}0.01211\\
\cAlignHack Group-VI \mbox{}\protect\newline Preventive regimen-1 &
  \cAlignHack 300 &
  \cAlignHack 15.65\ensuremath{\pm}0.1907 &
  \cAlignHack 6.20\ensuremath{\pm}0.02578 &
  \cAlignHack 0.51\ensuremath{\pm}0.01276\\
\cAlignHack Group-VII \mbox{}\protect\newline Preventive regimen-2 &
  \cAlignHack 600 &
  \cAlignHack 19.49\ensuremath{\pm}0.1922 &
  \cAlignHack 6.75\ensuremath{\pm}0.02569 &
  \cAlignHack 0.41\ensuremath{\pm}0.01276\\
\tblbottomrule 
\end{tabulary}\par 
\end{table*}

\bgroup
\fixFloatSize{images/37841891-a48d-4fca-8768-a0c12affb2c6-upicture1.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/37841891-a48d-4fca-8768-a0c12affb2c6-upicture1.png}{\includegraphics{images/37841891-a48d-4fca-8768-a0c12affb2c6-upicture1.png}}{}
\makeatother 
\caption{\boldmath {Urinary Excretion of Stone Forming Constituents in Control and Experimental Rats}}
\label{f-727414e0527e}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/bf92aeea-fa0d-4a80-afe8-5311cc713d80-upicture2.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/bf92aeea-fa0d-4a80-afe8-5311cc713d80-upicture2.png}{\includegraphics{images/bf92aeea-fa0d-4a80-afe8-5311cc713d80-upicture2.png}}{}
\makeatother 
\caption{\boldmath {Kidney Retention of Stone Forming Constituents in Control and Experimental Rats}}
\label{f-d2753aea8f07}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/c4800c2c-7326-45d2-a15c-2256ac198350-upicture3.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/c4800c2c-7326-45d2-a15c-2256ac198350-upicture3.png}{\includegraphics{images/c4800c2c-7326-45d2-a15c-2256ac198350-upicture3.png}}{}
\makeatother 
\caption{\boldmath {Serum Parameters in Control and Experimental Rats}}
\label{f-30d0f75af33f}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/27a389f5-391b-4509-94c6-62c038c42ae6-upicture4.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/27a389f5-391b-4509-94c6-62c038c42ae6-upicture4.png}{\includegraphics{images/27a389f5-391b-4509-94c6-62c038c42ae6-upicture4.png}}{}
\makeatother 
\caption{\boldmath {Urine Volume, Urine pH, Kidney Weight in Control and Experimental Rats}}
\label{f-579f8859bbec}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/ae6a3174-8194-4b21-b34c-5430996d6153-upicture5.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/ae6a3174-8194-4b21-b34c-5430996d6153-upicture5.png}{\includegraphics{images/ae6a3174-8194-4b21-b34c-5430996d6153-upicture5.png}}{}
\makeatother 
\caption{\boldmath {Section of Normal Control Rat Kidney Showing Normal Organization of Tubular Epithelial Cells and Glomeruli}}
\label{f-42a3afeabcda}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/de0f06e9-7430-4846-94c4-2a61e4c8c954-upicture6.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/de0f06e9-7430-4846-94c4-2a61e4c8c954-upicture6.png}{\includegraphics{images/de0f06e9-7430-4846-94c4-2a61e4c8c954-upicture6.png}}{}
\makeatother 
\caption{\boldmath {Section of Urolithic Control Rats Showing the Deposition of Crystals}}
\label{f-fcdcebcbaacd}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/6cacabdd-2565-4e59-9588-454338226a57-upicture7.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/6cacabdd-2565-4e59-9588-454338226a57-upicture7.png}{\includegraphics{images/6cacabdd-2565-4e59-9588-454338226a57-upicture7.png}}{}
\makeatother 
\caption{\boldmath {Section of Standard Showing Regenerative Changes in the Glomeruli and Tubules}}
\label{f-a2c0efdb9ee6}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/eed3c7b0-c627-4637-944f-7c7267998855-upicture8.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/eed3c7b0-c627-4637-944f-7c7267998855-upicture8.png}{\includegraphics{images/eed3c7b0-c627-4637-944f-7c7267998855-upicture8.png}}{}
\makeatother 
\caption{\boldmath {Section of Curative Regimens of Dose-1 and Dose-2 Showing Decrease in Crystal Deposition in the Tissues and Lesser Tissue Damage When Compared to Urolithic Control Rats}}
\label{f-64161c935d39}
\end{figure}
\egroup

\bgroup
\fixFloatSize{images/36bc14c5-b3bb-4dd2-86c2-317d603a9eba-upicture9.png}
\begin{figure}[!htbp]
\centering \makeatletter\IfFileExists{images/36bc14c5-b3bb-4dd2-86c2-317d603a9eba-upicture9.png}{\includegraphics{images/36bc14c5-b3bb-4dd2-86c2-317d603a9eba-upicture9.png}}{}
\makeatother 
\caption{\boldmath {Section of Preventive Regimens of Dose-1 and Dose-2 Showing Decreased Crystal Deposition in the Tissues and Lesser Tissue Damage when Compared to Urolithic Control Rats}}
\label{f-7f27def049a9}
\end{figure}
\egroup
\textbf{Evaluation of Poly-Herbal Suspension}

\textbf{Organoleptic Evaluation }

The color, odor and taste of the prepared Poly Herbal Suspension were noted.

\textbf{Physico-Chemical evaluation}

\textbf{Stability}

The stability studies were carried out for the formulation by storing the prepared poly-herbal suspension at different temperature conditions like 4\ensuremath{^{0}}C, 25\ensuremath{^{0}}C \& 45\ensuremath{^\circ}C for 1 week, then its stability was observed. During the stability studies no change in colour, odour, and taste was observed in the polyherbal suspension. And its redispersibility was also found to be good \unskip~\cite{1365009:24518645}.

\textbf{pH}

The pH has been established with the use of pH digitalized mtr. 30ml of polyherbal suspension was taken in beaker and dipped the bulb of the P\ensuremath{^{H}} meter into the formulation and the pH was measured.

\textbf{Viscosity}

The viscosity of polyherbal suspension\ensuremath{^{}} was determined by using digital Brookfield viscometer. 50ml of poly-herbal suspension was taken into 100ml beaker and the tip of viscometer was dipped into the beaker containing anti-urolithiatic suspension and its viscosity was measured \unskip~\cite{1365009:24518641}.

\textbf{Density}

The density of the prepared suspension was calculated.

\textbf{Sedimentation Volume}

The Sedimentation volume of the prepared formulation was calculated.

\textbf{Particle Size}

Calibrated the eye piece micrometer using the stage micrometer and calibration factor was calculated. The one fall like suspension has been positioned upon that slide and size of 25 particles was measured and calculated the average size of the particles.

\textbf{Redispersibility}

Possible problems have been evaluated through locating suspensions inside a 100 milliLitre measuring cyl. Now since collection but also sediment transport, a cyl has been turned and via 360\ensuremath{^{0}} about as 20 revolutions per minute. The top point is well taken whenever the bottom of a cyl is obvious like turbidity including that of the consistency like suspension particulate.

\textbf{Acute Oral Toxicity Study}

That whole process is followed through using oecd 423 (acute poisonous type method). A severe toxic type method is just a step-by-step process as for 3 living creatures of such a segregated by sex for every stage. Counting on its death rate as well as defunct condition of a living creatures and also the ordinary 2 to 3 stages could be essential to enable assessment upon that toxicity studies of a test compound. Such an methodology leads to its use amount of animals whereas letting as a appropriate information based sciences inference. The strategy was using outlined dose levels (2000, 1000, 500, 5 mg/kg body weight) and also the conclusions enable the one constituent to also be consistently rated but also categorized as per the global requirements process (ghs) is for categorization like chemical compounds that cause toxicity studies \unskip~\cite{1365009:24518652}.

\textbf{Experiments Test Methods}

Elderly wistar albino mice needs to weigh 160-180gm were being used for a research. It and beginning dose stage like PHf has been 200mg/kg body mass p. o~like most of the preparations contain ld50 valuation more of it than 200mg/kg p. o~dose quantity has been administered of about up overnight fasting conditions mice as for libitum. 

Nutrition has been decided to withhold further for three-four hours now since administering but also recognized as a signage such as toxic effect \unskip~\cite{1365009:24518647}.

\textbf{Slab Provoking Technique}

Ethylene glycol and ammonia solution caused urolithiasis: libitum connect of about normal food as well as drinkable water usually contains 0.75\% [v/v] ethylene glycol (eg) and 1\% [w/v] ammonia solution (ac) so that you can help in promoting hyperuricemia but also coax accumulation with in urinary tracts.

\textbf{Regular Drug}

Cystone 750 mg/kg body.weight.

\textbf{Animals}

Wister male mice (150-200g) have been acquired but were holed up inadequately regular laboratory situations but were supplied promotional rodent feed (Lipton India Ltd, Mumbai, India) but also room temperature water, libitum, the whole animal experimentation have been done as according nih regulations, now since receiving the permission of a institute's animals rights council (IAEC) like CPCSEA.

\textbf{Design of Experiments}

That whole study carried out over albino rats 150-200g, those who have been supplied with such a standard rodent eating plan (gold mohar, lipton india ltd., Calcutta) as well as liquid libitum. Already when one's use in research, the mice had been managed to keep through proper environmental situations, (temperature 25-28\ensuremath{^{0}}c and 12h light/dark cycle) [Table~\ref{tw-797a4804d7e5}]. Seven groups of six animals each were used. Group I~served as control, group II received the stone inducing agent, group III received the standard drug Cystone (p.o.). The remaining groups received the prepared suspension at different concentrations of 300mg/kg p.o and 600mg/kg p.o which were considered as preventive and curative regimens \unskip~\cite{1365009:24518644}.

\textbf{Urinary Evaluation}

There at finish of 28 days living creatures have been did weigh; holed up through physiologic enclosures such as 24 hours as well as urinary has been gathered. One fall like HCL has been go to add to a uric acid before becoming saved at 40\ensuremath{^{o}}C. Gathered samples have been approximated as a specifications including calcium, oxalate but also phosphorous \unskip~\cite{1365009:24518648}.

\textbf{Serum Analyze}

After experiment conducted, blood has been gathered first from retro-orbital below general anaesthetic situations but also living creatures have been made sacrifices along cervical decapitation. Serum has been kept separate besides centrifugation there as 10,000 rpm for 10 min as well as examined as a parametric like bun, creatine kinase and urea.

\textbf{Renal Homogenate Analyze}

A stomach has been sliced open of about start by removing both kidneys of each living creature. Isolated renal system have been kept clean out of extraneous tissue as well as maintained through 10\% neutral formalin. The kidneys have been left to dry at 80\ensuremath{^{o}}c inside a dried in an oven. A survey of 100mg of a dehydrated kidney has been heated through 10ml like 1n HCL just that 30 minute but also homogenized. The samples were centrifuged about as 2,000 rpm for 10 minutes and also the supernatant has been kept separate. That whole calcium, phosphate and oxalic acid material through kidney homogenate have been ascertained \unskip~\cite{1365009:24518639}.

\textbf{Histopathological Research}

Upon plasma draining, kidney sample was then have been purged as from control but also treated like living creature as well as cleaned to saline solution individually. Those that have been repaired through 10\% buffered formol just that 24hours. The formalin-fixed kidney specimens have been soaked as for haemotoxylin-eosin for photo microscopic observational data of kidney histopathologic architects, on such a carl zeiss binocular magnifying glass \unskip~\cite{1365009:24518649}.
    
\section{RESULTS AND DISCUSSION}
\textbf{Physico-chemical parameters}

\textbf{Organoleptic Evaluation }

The color, odor and taste of the prepared Poly Herbal Suspension were noted [Table~\ref{tw-a9acd22d1261} \& Table~\ref{tw-3505c5be94a4}].

\textbf{Physico-chemical Evaluation}

The pH, Viscosity, Density,\ensuremath{^{}}Sedimentation volume\ensuremath{^{, }}Particle size, Redispersibility [Table~\ref{tw-063b951f7bb0}].

\textbf{Moisture Content}

Moisture content of the above powered plants was performed and the results are presented in Table~\ref{tw-1f0d642d5821}.

\textbf{Ash Values}

Ash values of all the powered materials were performed and the results are presented in Table~\ref{tw-2a8c83f44440}.

\textbf{Extractive Values}

Extractive values of powered plant materials are presented in Table~\ref{tw-a3f602c422d1}.

Suspension formulation was prepared using combination of extracts of \textit{Tribulus terrestris} L. (Zygophyllaceae)\textit{, Aerva lanata} L. (Amaranthaceae)\textit{, Crataeva religiosa } Hook \& Frost. (Capparidaceae) and\textit{ Emblica officinalis} L.(Euphorbiaceae)\textit{. }

\textbf{Toxicity Results}

A body mass of both the mice during administering have been mentioned as well as stated there were no keeps changing through skin but also fur, eyes, mucosal membranes, blood circulation, autonomic but also nervous system and motor activity as well as behavior and then also recognized no sign of tremors, convulsions, salivary flow, diarrhoea, lethargy sleep as well as unconsciousness. The onset of toxicity and indications like toxic effects really aren't discovered. Further in research, Neither toxic effects and dying had been discovered at all these levels [Table~\ref{tw-eb00541d06fc}].

From the acute toxicity study, LD\ensuremath{_{50}} dose was found to be 2000mg/kg body weight for thepolyherbal suspension. The polyherbal suspension did not produce any significant behavioral changes. Hence 300 mg/kg body weight and 600mg/kg body weight was considered as therapeutic doses.

\textbf{Urine Analysis}

In the present study, chronic administration of 0.75\% [v/v] ethylene glycol (EG) and 1\% [w/v] ammonium chloride (AC) in drinking water to male Wistar rats resulted in hyperoxaluria. Oxalate, calcium and phosphate excretion were very high in calculi-induced animals [Figure~\ref{f-727414e0527e}]. However, the administration of polyherbal suspension significantly (p \textless\ 0.05) lowered the elevated levels of calcium, phosphate and oxalate in urine in a dose dependent manner in both curative and preventive regimen [Table~\ref{tw-36d0f6a25c6a}].

\textbf{Kidney Homogenate Analysis}

The same results were observed in kidney homogenate analysis also. The accumulation of the crystalline components in the renal tissue, namely oxalate, phosphate and calcium was increased in the stone forming rats. The herbal suspension significantly (p \textless\ 0.05) reduced the renal content of these stone forming constituents in both regimens [Table~\ref{tw-b2a49dfd5a12}]. Polyherbal suspension at 600mg/kg bodyweight has shown better activity than lower dose [Figure~\ref{f-d2753aea8f07}].

\textbf{Serum Analysis}

Serum analysis showed a marked increase of uric acid and BUN in calculi-induced animals. But the serum creatinine level was only slightly increased in group II, indicating marked renal damage [Table~\ref{tw-9d2a50d47447}]. The administration of polyherbal suspension significantly (p \textless\ 0.05) reduced the elevated serum levels of creatinine, uric acid and BUN in curative and preventive regimens [Figure~\ref{f-30d0f75af33f}].

\textbf{Urinary pH\ensuremath{^{}}}

The urinary pH in normal animals was between 6.0 and 7.0. Administration of 0.75\% ethylene glycol and 1\% ammonium chloride in drinking water to rats showed a significant reduction (p \textless\ 0.05) in urine pH [Figure~\ref{f-579f8859bbec}]. The urine pH of preventive and curative groups was between7-8 and it was comparable with control and standard groups [Table~\ref{tw-10940ab2f0f0}].

\textbf{Urine Volume}

Administration of 0.75\% ethylene glycol and 1\% ammonium chloride in drinking water to rats showed a significant (p \textless\ 0.05) reduction in urine volume [Figure~\ref{f-579f8859bbec}]. The urine volume of the preventive and curative groups treated with the prepared Poly herbal suspension was increased comparably with the control and the standard drug treated groups [Table~\ref{tw-10940ab2f0f0}].

\textbf{Kidney Weight}

Administration of 0.75\% ethylene glycol and 1\% ammonium chloride in drinking water to rats showed a significant (p \textless\ 0.05) increase in kidney weight which is supposed to be due to the deposition of stone forming constituents [Figure~\ref{f-579f8859bbec}]. The kidney weight of the preventive and curative groups treated with the prepared Poly herbal suspension were decreased comparably with the control and the standard drug treated groups which indicate that the prepared Poly herbal suspension significantly reduced the concentration of stone forming constituents [Table~\ref{tw-10940ab2f0f0}].

\textbf{Histopathology Results}

Histopathology results showed that administration of 0.75\% ethylene glycol and 1\% ammonium chloride in drinking water to rats caused deposition of crystals, but normal organization of tubular epithelial cells and glomeruli is observed in normal control rat kidney, the stone formation in the preventive and curative groups treated with the prepared Poly herbal suspension were decreased comparably with the control and the standard drug treated groups which indicate that the prepared Poly herbal suspension significantly reduced the concentration of stone forming constituents [Figure~\ref{f-42a3afeabcda}, Figure~\ref{f-fcdcebcbaacd}, Figure~\ref{f-a2c0efdb9ee6}, Figure~\ref{f-64161c935d39} \& Figure~\ref{f-7f27def049a9}].
    
\section{DISCUSSION}
Urolithiasis, stone formation in kidney, has ever been an annoying urinary disease for human and in India 11\% of the population is suffering with this urinary disorder. A urolithiasis patient, when seeks medical attention will be prescribed one or more of analgesics, anti-inflammatory and spasmolytic drugs by a physician. While lots of herbal therapies were used to overcome urolithiasis in the past, today mainly technological techniques like extracorporeal shock wave lithotripsy and percutaneous nephrolithotomy are used in urolithiasis treatment. However, it was reported that these updated techniques came up short in urolithiasis avoidance. In addition, some side effects such as hemorrhage, hypertension, tubercular necrosis and subsequent fibrosis of the kidney leading to cell injury and recurrence of renal stone formation were also reported. For this reason, alternative herbal therapies and techniques with fewer side effects should be researched.

The organic and inorganic constituents present in a drug or plant play a significant role in identification of crude drug. Physical constants like ash, extractive values and phytochemical analysis helps in establishing the pharmacopoeial standards of drug. The chief phytochemicals present in the different extracts were alkaloids, carbohydrates, glycosides, steroids, saponins, tannins, flavonoids, proteins, fixed oils and fats. These phytochemicals might be responsible for better activity of formulation.

In the current study, male rats were selected to induce urolithiasis because the urinary system of male rats resembles that of humans and also earlier studies have shown that the amount of stone depositing in female rats was significantly less. For this reason we have preferred to use this model to examine antiurolithiatic effect of Polyherbal suspension. In literature, there are lots of studies on urolithiasis model using ethylene glycol and ammonium chloride in different concentrations (0.4-2.5\%) and different usage times (7 days -1 month). In our experimental design, 1\% ethylene glycol and 1\% ammonium chloride solution was used as rat drinking water in urolithiasis groups for 28 days. Evidence in previous studies indicated that in response to 14 day period of ethylene glycol (0.75\% v/v) administration, young male albino rats form renal calculi composed mainly of calcium oxalate. Stone formation in ethylene glycol fed animals is caused by hyperoxaluria, which causes increased renal retention and excretion of oxalate. Similar results have been obtained when rats were treated with ethylene glycol and ammonium oxalate.

Obtained data showed that, oxalate and calcium excretion are progressively increased in calculi-induced animals (group 2.). Since it is accepted that hyperoxaluria is a far more significant risk factor in the pathogenesis of renal stones than hypercalciuria the changes in urinary oxalate levels are relatively much more important than these calcium. Increased urinary calcium is a factor favoring the nucleation and precipitation of calcium oxalate or calcium phosphate from urine and subsequent crystal growth. However, the Polyherbal suspension lowered the levels of oxalate as well as calcium excretion.

An increase in urinary phosphate is observed in calculi-induced rats (group 2). Increased urinary phosphate excretion along with oxalate stress seems to provide an environment appropriate for stone formation by forming calcium phosphate crystals, which epitaxially induces calcium oxalate deposition. Treatment with Poly herbal suspension restores phosphate levels, thus reducing the risk of stone formation.

The urine pH level was significantly lowered in polyherbal treated groups when compared to the calculi-induced group. In the present study serum concentrations of urea, uric acid and BUN were determined. The results showed a considerable increase in uric acid, creatinine and BUN in calculi-induced group than the intact control group and this may be due to the formation of stones in the urinary tract. The results are compatible with the present findings. On the other hand, the curative and prophylactic treatment with Poly herbal suspension significantly reduced the levels of uric acid. Creatinine and BUN when compared to urolithiasis group. It may be due to the /stone dissolving action of Polyherbal suspension or due to prevention of new stone formation in urinary system. The significant lowering of serum levels of accumulated waste product is attributed to the enhanced glomerular filtration rate.

From the histopathology results, it is clear that section of normal control rat kidney showed normal organization of tubular epithelial cells and glomeruli but the kidney tissue of urolithic control rats showed the deposition of crystals. Furthermore, tissue section of standard group showed regenerative changes in the glomeruli and tubules. Tissue section of preventive rats protected the tissue from damage and caused less/no crystal deposition. Curative regimens decreased the crystal deposition in the tissues and lower the tissue damage when compared to urolithic control rats. Similar histological observations earlier reported support the deposition of crystals and attainment of normalcy of tubular epithelial cells and glomeruli when treated with polyherbal suspension.
    
\section{CONCLUSION}
It is concluded that, scientific parameters based on phytochemical studies are essential in identifying and distinguishing the plant from others. Standardization of drugs and formulations of traditional system of medicine is of utmost importance to assure uniform quality and optimum biological efficacy. There was rigorous intention to find the effect of plant extracts in dissolution and prevention of the growth of urinary stones, So, aqueous extract of \textit{Aerva lanata} L., petroleum ether extract of \textit{Crataeva religiosa } Hook \& Frost., ethanol extract of \textit{Tribulus terrestris} L. and methanol extract of \textit{Emblica officinalis} L. were selected to prepare a polyherbal suspension and systematic work was carried out. From the studies, it was found that all the extracts of plants parts were effective in \textit{in-vivo} anti-urolithiatic activity on induced calcium oxalate crystals in rats and found noteworthy in treatment of renal calculosis. The present study indicates that the administration of polyherbal suspension to rat with ethylene glycol induced lithiasis reduced and prevented the growth of urinary stones, supporting folk information regarding anti urolithiatic activity of the plants. The polyherbal suspension resulted in an increase in urinary volume, decrease in calcium, oxalate and phosphate excretion. Further the histopathological examination of renal tissues showed drastic reduction in stone formation. So, it is concluded that the polyherbal suspension used inthe present study is an effective drug in the management and treatment of urolithiasis. However, further studies are required for authentication in human beings.
    
\section{ACKNOWLEDGEMENT }
The authors are thankful to Dr. Pavan Kumar Balagani Sir, Professor \& Principal, Gokula Krishna College of Phrmacy, Behind RTC Depot, NV Kandriga, Sullurpeta, SPSR Nellore Dist, Andhra Pradesh - 524121 providing the necessary facilities to carry out this work.

\textbf{Funding Support}

The author declares that no funding was supported for this study.

\textbf{Conflict of Interest}

The author declares there was no conflict of interest for this study.

\textbf{Contribution of Authors }

Authors declare that, the Research work by the names mentioned in the article and all the liabilities and claims related to the content of the article will be borne by the authors.
    

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