Regarding the book, Tlass has stated that "I intend through publication of this book to throw light on some secrets of the Jewish religion based on the conduct of the Jews and their fanaticism" and that both Eastern and Western civilizations threw Jews into ghettos only after recognizing their "destructive badness". He also claimed that since 1840, "...every mother warned her child: Do not stray far from home. The Jew may come by and put you in his sack to kill you and suck your blood for the Matzah of Zion."

Tlass was born in Rastan near the city of Homs to a prominent local Sunni Muslim family on 11 May 1932. His father, Abdul Qadir Tlass, was a minor Sunni notable who made a living during the Ottoman period by selling ammunition to the Turkish garrisons. On the other hand, members of his family also worked for the French occupiers after the First World War. His paternal grandmother was of Circassian origin and his mother was of Turkish descent. Tlass is said to also have some Alawite family connections through his mother. He received primary and secondary education in Homs. In 1952, he entered the Homs Military Academy.

Tlass married Lamia Al Jabiri in 1958. She is a member of the Aleppine aristocracy. His marriage secured his position among the traditional elite and enabled him to advance socially. They have four children: Nahid (born 1958), Firas (born 1960), Manaf (born 1964), and Sarya (born 1978). His daughter Nahid was married to Akram Ojjeh, who was a Saudi millionaire arms dealer. Nahid Tlass has lived in Paris since the onset of Syrian uprising. His younger daughter, Sarya, is married to a Lebanese from Baalbak.

During the 1960s, Hafez al-Assad rose to prominence in the Syrian government through the 1963 coup d'état, backed by the Ba'ath party. He then promoted Tlass to high-ranking military and party positions. In 1965, while he was Ba'athist army commander of Homs, Lieutenant Colonel Mustafa Tlass arrested his pro-government comrades. A 1966 coup by an Alawite-dominated Ba'ath faction further strengthened al-Assad, and by association Tlass. Tensions within the government soon became apparent, however, with al-Assad emerging as the prime proponent of a pragmatist, military-based faction opposed to the ideological radicalism of the dominant ultra-leftists. Syrian defeat in the 1967 Six Day War embarrassed the government, and in 1968 al-Assad managed to install Tlass as new Chief-of-Staff. After the debacle of an attempted Syrian intervention in the Black September conflict, the power struggle came to open conflict.

In 1969, Tlass led a military mission to Beijing, and secured weapons deals with the Chinese government. In a move deliberately calculated to antagonize the Soviet Union to stay out of the succession dispute then going on in Syria, Mustafa Tlass allowed himself to be photographed waving Mao Zedong's Little Red Book, just two months after bloody clashes between Chinese and Soviet armies on the Ussuri river. The Soviet Union then agreed to back down and sell Syria weapons.

Under cover of the 1970 "Corrective Revolution", Hafez al-Assad seized power and installed himself as Dictator. Tlass was promoted to minister of defense in 1972, and became one of al-Assad's most trusted loyalists during the following 30 years of one-man rule in Syria. As'ad AbuKhalil argues that Mustafa Tlass was well-suited for Hafez al-Assad as a defense minister in that "he had no power base, he was mediocre, and he had no political skills, and his loyalty to his boss was complete." During his term as defense minister, Mustafa Tlass was functional in suppressing all dissent regardless of being Islamists or democrats.

There have been three missing Israeli Soldiers in the Bekaa valley since the June 1982 war in Lebanon. Tlass allegedly told a Saudi magazine: "We sent Israel the bones of dogs, and Israel may protest as much as it likes."

In 1983, Tlass wrote and published The Matzah of Zion, which is a treatment of the Damascus affair of 1840 that repeats the ancient "blood libel", that Jews use the blood of murdered non-Jews in religious rituals such as baking Matza bread. In this book, he argues that the true religious beliefs of Jews are "black hatred against all humans and religions," and that no Arab country should ever sign a peace treaty with Israel. Tlass re-printed the book several times, and stands by its conclusions. Following the book's publication, Tlass told Der Spiegel, that this accusation against Jews was valid and that his book is "an historical study ... based on documents from France, Vienna and the American University in Beirut."

In 1991 The Matzah of Zion was translated into English. Egyptian Producer Munir Radhi subsequently decided it was the ideal "Arab answer" to the film Schindler's List and later announced plans to produce a film adaptation of The Matzah of Zion. The book also reportedly served as a "scientific" basis for a renewal of the blood libel charge in international forums. In 2001, Al-Ahram published an article titled "A Jewish Matzah Made from Arab Blood" which summarized The Matzah of Zion, concluding that: "The bestial drive to knead Passover matzahs with the blood of non-Jews is [confirmed] in the records of the Palestinian police where there are many recorded cases of the bodies of Arab children who had disappeared being found, torn to pieces without a single drop of blood. The most reasonable explanation is that the blood was taken to be kneaded into the dough of extremist Jews to be used in matzahs to be devoured during Passover."

In 1998, Syrian Defense Minister Tlass boasted to Al Bayan newspaper that he was the one who gave the green light to "the resistance" in Lebanon to attack and kill 241 US marines and 58 French paratroopers, but that he prevented attacks on the Italian Soldiers of the multi-national force because "I do not want a single tear falling from the eyes of [Italian actress] Gina Lollobrigida, whom [I] loved ever since my youth." In October of the same year, Tlass stated that there was no such country as Jordan, but only "South Syria".

On 19 October 1999, defence minister of China, General Chi Haotian, after meeting with Mustafa Tlass in Damascus to discuss expanding military ties between Syria and China, flew directly to Israel and met with Ehud Barak, the then prime minister and defence minister of Israel where they discussed military relations. Among the military arrangements was a 1 billion dollar Israeli Russian sale of military aircraft to China, which were to be jointly produced by Russia and Israel.

The succession of Bashar al-Assad, Hafez's son, seems to have been secured by a group of senior officials, including Tlass. After the death of Assad in 2000, a 9-member committee was formed to oversee the transition period, and Tlass was among its members.

Whether true or not, Tlass and his supporters were viewed by many as opponents of the discreet liberalization pursued by the younger al-Assad, and to maintain Syria's hardline foreign policy stances; but also as fighting for established privileges, having been heavily involved in government corruption. In February 2002 in the Jordanian daily Al Dustour stated that Tlass submitted his letter of resignation to Bashar al-Assad, and was set to step down in July 2002. However, in 2004, Tlass was replaced by Hasan Turkmani as defense minister. It is also argued that Shawkat pushed for the removal of Mustafa Tlass. Tlass also quit the regional command in 2005.

In an interview which aired RT on 8 June 2009 (as translated by MEMRI), Tlass claimed that Actress Gina Lollobrigida had once told him that he was the "one love in my life." He also claimed that Lady Diana wrote him letters that "were full of love and appreciation." He also claimed that Prince Charles gave him a submachine gun as a gift.

Mustafa Tlass and his son, Firas, both left Syria after the revolt against Assad began in 2011. Mustafa Tlass left for France for what he described as medical treatment. Firas, a Business tycoon, left Syria for Egypt in 2011, too. It is also reported that he is in Dubai.

In July 2012, Manaf Tlass, a Syrian officer and another son of Mustafa, defected from the Assad government and fled to Turkey and then to France.

Tlass died on 27 June 2017 in Avicenne Hospital in Paris, France at the age of 85.

Tlass also wrote books about Syria’s military and political history and also books of poetry, general Arab history, and a history of the military tactics used by Muhammad. His writings allegedly reflect anti-Semitism and belief in conspiracy theories. He also published two-volume memoirs, namely Mirat Hayati (Reflections of my life) in 2005.